Modified release compositions comprising tacrolimus

ABSTRACT

A modified release composition comprising tacrolimus releases less than 20% w/w of the active ingredient within 0.5 hours when subjected to an in vitro dissolution test using USP Paddle method and using 0.1 N HCl as dissolution medium and has increased bioavailability by effectively reducing or even avoiding the effects of CYP3A4 metabolism. The modified composition may be coated with an enteric coating; and/or may comprise a solid dispersion or a solid solution of tacrolimus in a hydrophilic or water-miscible vehicle and one or more modifying release agents; and/or may comprise a solid dispersion or a solid solution of tacrolimus in an amphiphilic or hydrophobic vehicle and optionally one or more modifying release agents.

This application is a continuation of U.S. patent application Ser. No.10/569,862, filed Feb. 27, 2006, which is the U.S. national phase ofInternational Application No. PCT/DK2004/000573, filed Aug. 30, 2004,which claims the benefit of U.S. Provisional Application No. 60/529,793(filed Dec. 15, 2003) and Danish Patent Application Nos. PA 2003-01232(filed Aug. 29, 2003), PA 2003-01837 (filed Dec. 11, 2003), PA2004-00079 (filed Jan. 21, 2004), PA 2004-00463 (filed Mar. 23, 2004),and PA 2004-00467 (filed Mar. 23, 2004), each of which is incorporatedherein by reference in its entirety.

The present invention relates to a pharmaceutical composition and/ordosage forms, preferably oral unit dosage forms, comprising tacrolimusor an analogue thereof having a modified release profiles when subjectedto a conventional dissolution method, which is believed to reflect theactual rate and timing of release of active ingredient in vivo, thenovel composition effectively reducing or even avoiding the effects ofCYP3A4 metabolism.

BACKGROUND OF THE INVENTION

Tacrolimus, also known as FK-506 or FR-900506, has the chemicaltricyclic structure shown below:

corresponding to C₄₄H₆₉NO₁₂. Tacrolimus appears in the form of whitecrystals or crystalline powder. It is practically insoluble in water,freely soluble in ethanol and very soluble in methanol and chloroform.

The preparation of tacrolimus is described in EP-A-0 184 162 andanalogues of tacrolimus are disclosed e.g. in EP-A-0 444 659 and U.S.Pat. No. 6,387,918, which are both hereby incorporated by reference.

Tacrolimus is a macrolide compound with useful immunosuppressiveactivity, antimicrobial activity and other pharmacological activitiesand is of value for the treatment or prevention of rejection reactionsby transplantation of organs or tissues, graft versus host diseases,autoimmune diseases and infectious diseases. Tacrolimus prolongs thesurvival of the host and transplanted graft in animal transplant modelsof liver, kidney, heart, bone marrow and small bowel and pancreas, lungand trachea, skin, cornea and limb.

In animals, tacrolimus has been demonstrated to suppress some humoralimmunity and, to a greater extent, cell-mediated reactions such asallograft rejection, delayed type hypersensitivity, collagen-inducedarthritis, experimental allergic encephalomyelitis and graft-versus-hostdisease.

Tacrolimus inhibits T-lymphocyte activation, although the exactmechanism of action is unknown. Experimental evidence suggest thattacrolimus binds to an intracellular protein, FKBP-12. A complex oftacrolimus-FKBP-12, calcium, calmodulin, and calcineurin is then formedand the phosphatase activity of calcineurin inhibited. This effect mayprevent the dephosphorylation and translocation of nuclear factor ofactivated T-cells, a nuclear component thought to initiate genetranscription for the formation of lymphokines. The net result is theinhibition of T-lymphocyte activation, i.e. immunosuppression.

Tacrolimus is extensively metabolized by the CYP3A4 isoenzyme in the gutwall and liver. Therefore, drugs that affect this isoenzyme mayinfluence absorption and the subsequent elimination of systemicallyabsorbed tacrolimus. Inhibitors of CYP3A4 may increase tacrolimuslevels, while inducers of CYP3A4 may increase the metabolism oftacrolimus and decrease tacrolimus levels. Accordingly, tacrolimus maybe administered together with one or more CYP3A4 inhibitors in order toimprove the overall bioavailability.

Usually tacrolimus is administered orally and is therefore absorbed fromthe gastrointestinal tract. It has been observed that the absorption isnegatively influenced by the simultaneous ingestion of food. Thus, therate and extent of tacrolimus absorption were greatest under fastedconditions.

In general, it is known that the absorption and bioavailability of atherapeutically active substance can be affected by a variety of factorswhen administered orally. Such factors include the presence of food inthe gastrointestinal tract and, in general, the gastric residence timeof a drug substance is significantly longer in the presence of food thanin the fasted state. If the bioavailability of a drug substance isaffected beyond a certain point due to the presence of food in thegastrointestinal tract, the drug substance is said to exhibit a foodeffect. Food effects are important because absorption and hence theplasma levels becomes highly variable depending on food intake.Absorption into the bloodstream may be adversely affected to the pointthat the patient risks insufficient absorption to remedy the conditionfor which the drug was administered. On the other hand, the very highpeak concentrations seen at fasted conditions occasionally, may verywell induce significant side effects, of nephro- or neuro-toxic origin,as well as GI side-effects and others.

Absorption of tacrolimus from the gastrointestinal tract after oraladministration is rapid with a mean time-to-peak concentration (t_(max))of approximately 1-2 hours after administration to healthy subjects orkidney or liver transplanted patients, but incomplete and variable. Thebioavailability is generally as low as at the most about 20% after oraladministration.

Frequently observed side effects are vomiting and nausea but sideeffects like tremor, headache, hypertension, renal dysfunction,hyperkalemia, hypomagnesaemia, hyperglycemia, insomnia, diarrhea,constipation, abdominal pain, nephrotoxicity and neurotoxicity are alsoobserved.

For oral administration, tacrolimus is currently formulated and marketedas soft gelatine capsules comprising the equivalent of 0.5, 1 or 5 mganhydrous tacrolimus and marketed under the trade name Prograf® andProtropic®. The recommended initial oral dose is from about 0.1 to 0.2mg/kg/day in patients. The dose aims at a certain trough plasma levelfrom about 5 to about 20 ng/ml. Prograf® is indicated for theprophylaxis of organ rejection in patients receiving allogeneic liver orkidney transplants.

There remains a need for novel pharmaceutical compositions and/or dosageforms comprising tacrolimus exhibiting enhanced bioavailability. Anincreased bioavailability may allow a reduction in the dosage unitstaken by a patient, e.g. down to a single dose daily, and may alsoreduce or negate the need for food to be takes simultaneously with thedosage form thereby allowing patients more freedom on when the drug istaken. Furthermore, it is contemplated that fluctuations in the plasmaconcentration versus time profile may be significantly reduced. Further,enhanced bioavailability may also result in a more reproducible (i.e.less variable compared to that of Prograf®) release profile.

BRIEF SUMMARY OF THE INVENTION

The inventors have found that the bioavailability of tacrolimus issignificantly increased when tacrolimus is administered to a mammal in amodified or controlled release composition providing a rate and a timingof release of active ingredient, i.e. an in vivo release profile,effectively reducing or even avoiding the effects of CYP3A4 metabolism.

It is believed that conventional in vitro dissolution methods correlateto or at least reflect the actual in vivo modified release profile inman. In accordance herewith, the present invention provides, in itsfirst aspect, a solid pharmaceutical composition comprising an activeingredient selected among tacrolimus and analogues thereof, wherein lessthan 20% w/w of the active ingredient is released within 0.5 hours, whensubjected to an in vitro dissolution test using USP Paddle method andusing 0.1 N HCl as dissolution medium. This modified release profile isobtained by providing a pharmaceutical composition which

-   -   i) is coated with an enteric coating; and/or    -   ii) comprises a solid dispersion or, preferably, a solid        solution of active ingredient, i.e. tacrolimus or an analogue        thereof, in a hydrophilic or water-miscible vehicle and one or        more modifying release agents; and/or    -   iii) comprises a solid dispersion or, preferably, a solid        solution of active ingredient, i.e. tacrolimus or an analogue        thereof, in an amphiphilic or hydrophobic vehicle and optionally        one or more modifying release agents.

In a further aspect, the invention relates to solid dosage forms,especially oral dosage forms, comprising the composition of theinvention, the solid dosage forms exhibiting a modified release profile.Delaying the release of tacrolimus to the distal part of duodenum mayreduce the drug related gastro-intestinal related side effects and therelatively high degree of metabolism in the proximal part of thegastrointestinal tract (CYP3A4 mediated metabolism). This can be donewithout loosing systemic bioavailability due to the unique compositionsof the invention, preferably compositions comprising the activeingredient fully or partly dissolved in a vehicle to form a soliddispersion and/or a solid solution at ambient temperature.

In yet further aspects, the invention relates to use of the presentpharmaceutical composition to enhance the oral bioavailability oftacrolimus, to use of the present composition in the preparation ofmedicines or medicaments, especially in the preparation of useful soliddosage forms.

DETAILED DESCRIPTION OF THE INVENTION Definitions

As used herein, the term “active ingredient” or “active pharmaceuticalingredient” means any component that is intended to furnishpharmacological activity or other direct effect in the diagnosis, cure,mitigation, treatment, or prevention of disease, or to affect thestructure or any function of the body of man or other animals. The termincludes those components that may undergo chemical change in themanufacture of the drug product and are present in the drug product in amodified form intended to furnish the specified activity or effect.

In the present context, the term “hydrophilic” describes that something‘likes water’, i.e. a hydrophilic molecule or portion of a molecule isone that typically is electrically polarized and capable of forminghydrogen bonds with water molecules, enabling it dissolve more readilyin water than in oil or other “non-polar” solvents.

In the present context, the term “amphiphilic” describes a molecule (asa surfactant) having a polar water-soluble group attached to awater-insoluble hydrocarbon chain. Thus, one end of the molecule ishydrophilic (polar) and the other is hydrophobic (non-polar).

In the present context, the term “hydrophobic” denotes a compoundtending to be electrically neutral and non-polar, and thus preferringother neutral and nonpolar solvents or molecular environments.

As used herein, the term “vehicle” means any solvent or carrier fluid ina pharmaceutical product that has no pharmacological role. For example,water is the vehicle for xilocaine and propylene glycol is the vehiclefor many antibiotics.

In the present context, the term “solid dispersion” denotes a drug oractive ingredient or substance dispersed on a particulate level in aninert vehicle, carrier, diluent or matrix in the solid state, i.e.usually a fine particulate dispersion.

In the present context, the term “solid solution” denotes a drug oractive ingredient or substance dissolved on a molecular level in aninert vehicle, carrier, diluent or matrix in the solid state.

As used herein, the term “analogue” means a chemical compound that isstructurally similar to another.

The term “drug” means a compound intended for use in diagnosis, cure,mitigation, treatment, or prevention of disease in man or other animals.

In this context, the term “dosage form” means the form in which the drugis delivered to the patient. This could be parenteral, topical, tablet,oral (liquid or dissolved powder), suppository, inhalation, transdermal,etc.

As used herein, the term “bioavailability” denotes the degree means towhich a drug or other substance becomes available to the target tissueafter administration.

As used herein, the term “bioequivalency” denotes a scientific basis onwhich generic and brand name drugs are compared with one another. Forexample, drugs are bioequivalent if they enter circulation at the samerate when given in similar doses under similar conditions. Parametersoften used in bioequivalence studies are t_(max), c_(max),AUC_(0-Infinity), AUC_(0-t). Other relevant parameters may be W₅₀, W₇₅and/or MRT. Accordingly, at least one of these parameters may be appliedwhen determining whether bioequivalence is present. Furthermore, in thepresent context, two compositions are regarded as bioequivalent if thevalue of the parameter used is within 80-125% of that of Prograf® or asimilar commercially available tacrolimus-containing product used in thetest.

In the present context “t_(max)” denotes the time to reach the maximalplasma concentration (c_(max)) after administration; AUC_(0-Infinity)denotes the area under the plasma concentration versus time curve fromtime 0 to infinity; AUC_(0-t) denotes the area under the plasmaconcentration versus time curve from time 0 to time t; W₅₀ denotes thetime where the plasma concentration is 50% or more of C_(max); W₇₅denotes the time where the plasma concentration is 75% or more ofC_(max); and MRT denotes mean residence time for tacrolimus (and/or ananalogue thereof).

In this context, the term “medicine” means a compound used to treatdisease, injury or pain. Medicine is justly distributed into“prophylactic,” i.e. the art of preserving health, and “therapeutic”,i.e. the art of restoring health.

In the present context, the terms “controlled release” and “modifiedrelease” are intended to be equivalent terms covering any type ofrelease of tacrolimus from a composition of the invention that isappropriate to obtain a specific therapeutic or prophylactic responseafter administration to a subject. A person skilled in the art knows howcontrolled release/modified release differs from the release of plaintablets or capsules. The terms “release in a controlled manner” or“release in a modified manner” have the same meaning as stated above.The terms include slow release (that results in a lower C_(max) andlater t_(max), but t % is unchanged), extended release (that results ina lower C_(max), later t_(max), but apparent t_(1/2) is longer); delayedrelease (that result in an unchanged C_(max), but lag time and,accordingly, t_(max) is delayed, and t_(1/2) is unchanged) as well aspulsatile release, burst release, sustained release, prolonged release,chrono-optimized release, fast release (to obtain an enhanced onset ofaction) etc. Included in the terms is also e.g. utilization of specificconditions within the body e.g. different enzymes or pH changes in orderto control the release of the drug substance.

In this context, the term “erosion” or “eroding” means a gradualbreakdown of the surface of a material or structure, for example of atablet or the coating of a tablet.

The present invention provides pharmaceutical compositions and soliddosage forms for improved treatment of conditions that respond totacrolimus treatment, especially compositions and dosage forms providingmodified release of the active ingredient in order to enhance thebioavailability thereof.

The active ingredient in the inventive compositions is preferablytacrolimus or any analogue or derivative of tacrolimus, which exhibitseither a pharmacological or a therapeutical activity, which is at leastequivalent to that of tacrolimus (FK-506 or FR-900506). However, withinthe scope of the present invention is tacrolimus in any physical form(crystals, amorphous powder, any possible polymorphs, any possiblesolvates including the hydrate, anhydrate, complexes thereof etc.).Included is also any analogue, derivative or active metabolite oftacrolimus, pharmaceutically acceptable salts, solvates, complexes andprodrugs thereof.

Thus, in a preferred embodiment, the present invention provides a solidpharmaceutical composition comprising an active ingredient selectedamong tacrolimus and analogues thereof, wherein less than 20% w/w of theactive ingredient is released within 0.5 hours, when subjected to an invitro dissolution test using USP Paddle method and using 0.1 N HCl asdissolution medium; preferably wherein less than 20% w/w, morepreferably less than 10% w/w of the active ingredient is released within3 hours.

It is believed that such a release profile significantly enhances thebioavailability of tacrolimus in mammals, since all or a major part ofthe active ingredient is in fact released in the gastrointestinal tractin such as manner that CYP3A4 metabolism is substantially avoided or atleast significantly reduced. Further, it is contemplated that thiseffect is correlated to or at least reflected to the in vitrodissolution profile of the solid pharmaceutical composition and/ordosage forms of the invention, which profile is easily found whensubjecting the composition and/or dosage form to a conventional in vitrodissolution method according to e.g. USP. It is believed that any USP invitro dissolution method is useful for the present purpose.

For example, the solid pharmaceutical composition of the inventionreleases at least 50% w/w of the active ingredient within 4 hours,preferably within 2.5 hours, when subjected to an in vitro dissolutiontest using USP Paddle method and using 0.1N HCl as dissolution mediumduring the first 2 hours and then using a dissolution medium having a pHof 6.8.

Using a less conventional dissolution medium, the composition of theinvention releases less than 50 w/w %, especially less than 40 w/w %, ofthe active ingredient within 8 hours, preferably within 15 hours, whensubjected to an in vitro dissolution test using USP Paddle method and anaqueous dissolution medium adjusted to pH 4.5 with 0.005%hydroxypropylcellulose.

The desired modified release profile of the pharmaceutical compositionmay be provided by

-   -   i) coating the composition with an enteric coating; and/or    -   ii) using a pharmaceutical composition comprising a solid        dispersion or solid solution of active ingredient, i.e.        tacrolimus or an analogue thereof, in a hydrophilic or        water-miscible vehicle and one or more modifying release agents;        and/or    -   iii) using a pharmaceutical composition comprising a solid        dispersion or solid solution of active ingredient, i.e.        tacrolimus or an analogue thereof, in a hydrophobic vehicle and        optionally one or more modifying release agents.

In one embodiment of the invention, there is provided a modified releasetacrolimus-containing pharmaceutical composition which is entero-coatedas described herein.

In another embodiment of the invention, there is provided a modifiedrelease tacrolimus-containing pharmaceutical composition having theactive ingredient dissolved or dispersed in a hydrophobic vehicle asdescribed herein, preferably in an oil, an oily material, a wax or afatty acid derivative, more preferably a wax having a low melting pointsuch as for example glyceryl monostearate.

In yet another embodiment of the invention, there is provided a modifiedrelease tacrolimus-containing pharmaceutical composition having theactive ingredient dissolved or dispersed in a hydrophilic orwater-miscible vehicle as described herein, preferably a vehicleselected among polyethylene glycols, polyoxyethylene oxides, poloxamers,polyoxyethylene stearates, poly-epsilon caprolactone, polyglycolizedglycerides such as Gelucire®, and mixtures thereof, more preferablypolyethylene glycol optionally in mixture with a poloxamer. A specificexample of a useful mixture is a mixture of 70 w/w % polyethylene glycol6000 (PEG6000) and 30 w/w % poloxamer 188.

In a further aspect, the present invention relates to a pharmaceuticalcomposition in particulate form comprising tacrolimus and/or an analoguethereof together with one or more pharmaceutically acceptableexcipients, wherein the composition upon oral administration to a mammalin need thereof exhibits an AUC/AUC_(Prograf®) value of at least about1.3, the AUC values being determined under similar conditions.

As it appears from the examples herein the bioavailability obtainedafter administration of a composition according to the invention ismarkedly improved. Thus, in specific embodiments, the AUC/AUC_(Prograf®)value is at least about 1.5 such as about 1.75 or more, about 1.8 ormore, about 1.9 or more, about 2.0 or more, about 2.5 or more, about2.75 or more, about 3.0 or more, about 3.25 or more, about 3.5 or more,about 3.75 or more, about 4.0 or more, about 4.25 or more, about 4.5 ormore, about 4.75 or more or about 5.0 or more, the AUC values beingdetermined under similar conditions.

After oral administration of a pharmaceutical composition according tothe present invention it is contemplated that the plasma concentrationversus time profile show an extended period of time in which the plasmaconcentration is maintained within the therapeutic window (i.e. theplasma concentration leads to a therapeutic effect) without leading toserious unwanted side effects. Thus, a reduction in peak concentrationis also observed. Accordingly, the invention relates to a pharmaceuticalcomposition in particulate form comprising tacrolimus together with oneor more pharmaceutically acceptable excipient, wherein the compositionupon oral administration to a mammal in need thereof releases tacrolimusin a controlled manner and exhibits a C_(max) that is at the most about80% of that of C_(max) for Prograf® tablets such as, e.g., at the mostabout 75%, at the most about 70%, at the most about 65%, at the mostabout 60%, at the most about 55%, at the most about 50%, at the mostabout 45% or at the most about 40%.

In the present context the terms controlled release and modified releaseare intended to be equivalent terms covering any type of release oftacrolimus from a composition of the invention that is appropriate toobtain a specific therapeutic or prophylactic response afteradministration to a subject. A person skilled in the art knows howcontrolled release/modified release differs from the release of plaintablets or capsules. The terms “release in a controlled manner” or“release in a modified manner” have the same meaning as stated above.

The terms controlled release/modified release include slow release (thatresults in a lower C_(max) and later t_(max), but t_(1/2) is unchanged),extended release (that results in a lower C_(max), later t_(max), butapparent t_(1/2) is longer); delayed release (that result in anunchanged C_(max), but lag time and, accordingly, t_(max) is delayed,and t_(1/2) is unchanged) as well as pulsatile release, burst release,sustained release, prolonged release, chrono-optimized release, fastrelease (to obtain an enhanced onset of action) etc. Included in theterms is also e.g. utilization of specific conditions within the bodye.g. different enzymes or pH changes in order to control the release ofthe drug substance.

To be more specific, after oral administration to a mammal, including ahuman, of a pharmaceutical composition according to the presentinvention containing a dose of 5 mg tacrolimus, tacrolimus is releasedin a controlled manner and will exhibit a C_(max) that is at the mostabout 30 ng/ml such as, e.g. at the most about 25 ng/ml or at the mostabout 20 ng/ml.

However, a reduction in peak concentration may not lead to a decrease intherapeutic effect as long as the plasma concentration of tacrolimus ismaintained within the therapeutic window. Accordingly, the presentinvention also relates to a pharmaceutical composition, wherein W₅₀ isat least about 2 hours, such as, e.g., at least about 3 hours, at leastabout 4 hours, at least about 5 hours, at least about 6 hours, at leastabout 7 hours, at least about 8 hours, at least about 9 hours, about 10hours or more, about 11 hours or more, about 12 hours or more, about 13hours or about 14 hours or more.

Furthermore or moreover, a composition according to the invention has aC_(diff)=[C_(max)−C_(t) (t=12 hours)] that is less than that of Prograf®under the same conditions. If C_(diff) for Prograf® is set to 100 thenC_(diff) of a composition according to the invention is normally 90 orless such as, e.g., about 85 or less, about 80 or less, about 75 orless, about 70 or less, about 65 or less, about 60 or less, about 55 orless, about 50 or less, about 45 or less or about 40 or less.

More specifically, after oral administration to a mammal, including ahuman, of a pharmaceutical composition of the invention containing 5 mgof tacrolimus, tacrolimus is released in a controlled manner andexhibits a C_(diff) of about 20 ng/ml or less such as, e.g., about 15ng/ml or less, about 13 ng/ml or less or about 10 ng/ml or less.

A pharmaceutical composition according to the invention releasestacrolimus in a controlled manner in order to extend the therapeuticaction of tacrolimus. In one aspect the release may be pH dependant,i.e. the release predominantly takes place after passage of the stomach.Such a pH dependent release is mainly provided by means of entericcoating material as described herein. The release may also be pHindependent, e.g. by providing the composition with a controlled releasecoating such as, e.g. a cellulose based coating like e.g. ethylcelluloseor by providing the composition in the form of a matrix composition suchas, e.g., a hydrophilic cellulose polymer matrix type e.g. based onHPMC. A combination may of course also be employed.

In general, the change in bioavailability and/or the changes in otherbioavailability related parameters are normally determined by in vivostudies in a suitable animal model testing the compositions in questiontogether with e.g. Prograf® or a similar commercially availabletacrolimus-containing product. The use of a dog model for establishingevidence of the bioavailability of certain formulations is generalpractice in the pharmaceutical industry.

The studies relevant for tacrolimus are non-randomized, cross-overstudies, where each dog is it's own control. Four dogs, and fourtreatments are normally applied. As no iv injections are given, thebioavailabilities obtained are relative.

Further it has surprisingly been found that the need for simultaneousfood intake in order to secure a sufficient uptake of tacrolimus issignificantly reduced or even completely abolished.

Thus, the pharmaceutical compositions according to the invention providesignificant higher bioavailability of tacrolimus, which may reduce thenumber of daily administered dosage units, and reduce or abolish theneed for administration in connection with food intake, which providefor a higher degree of freedom for the recipient of the pharmaceuticalcompositions, and consequently the patients acceptance and/or compliancemay be significantly improved. Furthermore, the compositions provide asignificant reduction in side effects, especially side effect related toa high peak concentration (such as, e.g., nephro- and neuro-toxicity,diarrhea, constipation, abdominal pain, nausea etc) and provide for anextended release of tacrolimus leading to a better therapy.

As mentioned above, one of the major challenges with respect toformulation of tacrolimus compositions is to avoid an adverse foodeffect. In general, tacrolimus is much better absorbed when it isadministered orally without food. A great variation in bioavailabilityis therefore seen following administration with or without food. Thisdependency makes it difficult to give precise guidelines as to how largea dose that should be administered and, furthermore, it requiresinformation to the patient about the dosing regime. The presentinvention aims at providing compositions wherein the adverse food effectis reduced. Thus, the present invention provides a composition, whichdoes not exhibit a significant adverse food effect after administrationof the composition to a mammal in need of such a treatment as evidencedby a value of (AUC_(fed)/AUC_(fasted)) of at least about 0.85 with alower 90% confidence limit of at least 0.75.

More specifically, a pharmaceutical composition according to theinvention has a value of (AUC_(fed)/AUC_(fasted)) of about 0.9 or moresuch as, e.g., about 0.95 or more, about 0.97 or more or about 1 or moresuch as, e.g., up to about 1.1 or up to about 1.2.

A further advantage of a composition of the present invention is thepossibility of obtaining an effective therapeutic response with adecreased dosage compared to traditional oral treatment. Accordingly,upon oral administration to a mammal in need thereof a pharmaceuticalcomposition according to the invention releases tacrolimus or ananalogue thereof in a controlled manner and the composition isessentially bioequivalent with Prograf® or a similar commerciallyavailable tacrolimus-containing product when administered in a dosisthat is at the about most about 85% w/w such as, e.g., at the most about80% w/w, at the most about 75%, at the most about 70% w/w, at the mostabout 65% w/w, at the most about 60% w/w, at the most about 55% w/w orat the most about 50% w/w of the dose of tacrolimus administered in theform of Prograf® or a similar commercially availabletacrolimus-containing product.

Parameters often used in bioequivalence studies are t_(max), c_(max),AUC_(0-infinity), AUC_(0-t). Other relevant parameters may be W₅₀, W₇₅and/or MRT. Accordingly, at least one of these parameters may be appliedwhen determining whether bioequivalence is present. Furthermore, in thepresent context, two compositions are regarded as bioequivalent if valueof the parameter used is within 80-125% of that of Prograf® or a similarcommercially available tacrolimus-containing product used in the test.

In the present context “t_(max)” denotes the time to reach the maximalplasma concentration (c_(max)) after administration; AUC_(0-infinity)denotes the area under the plasma concentration versus time curve fromtime 0 to infinity; AUC_(0-t) denotes the area under the plasmaconcentration versus time curve from time 0 to time t; W₅₀ denotes thetime where the plasma concentration is 50% or more of C_(max); W₇₅denotes the time where the plasma concentration is 75% or more ofC_(max); and MRT denotes mean residence time for tacrolimus (and/or ananalogue thereof).

Two other main disadvantages associated with treatment or prophylaxiswith tacrolimus is the relative high incidence of side effects and arelatively high inter-individual variation. It is envisaged that acomposition according to the invention will lead to a reduction in sideeffects. The reduction may be in terms of reduced frequency or in termsof severity. The side effects in question include e.g. nephro- andneuro-toxicity, diarrhea, constipation, abdominal pain, nausea etc. Inone aspect the invention concerns a pharmaceutical composition inparticulate form comprising tacrolimus or an analogue thereof togetherwith one or more pharmaceutically acceptable excipient, wherein thecomposition upon oral administration to a mammal in need thereofreleases tacrolimus or an analogue thereof in a controlled manner andreduces side effects compared to those of Prograf® administered underthe same conditions and in a dose that provides an equivalenttherapeutic effect.

Increasing the bioavailability, the Area Under the Curve, will normallyreduce the intra- and inter-variability related to absorption of a drugsubstance. This is particularly true; whenever the low and impairedbioavailability is a consequence of poor water solubility. It iscontemplated that compositions according to the invention will provide aCV (Coefficient of Variation) on Area under Curve data that aresignificantly smaller than with Prograf® and like products.

As mentioned hereinbefore, one of the basic features of the presentinvention is that it is possible to obtain an improvement in thebioavailability by oral administration of a composition of the presentinvention. Normally, a low bioavailability of a drug substance afteroral administration is a barrier for design of a controlled or modifiedrelease composition of the drug substance due to the fact that it isalmost impossible to obtain effective drug levels over a prolongedperiod of time. However, with the present technology it is possible toobtain a significantly improved bioavailability and thereby possible todesign controlled, modified or delayed release compositions.

Tacrolimus is extensively metabolized by the CYP3A4 isoenzyme in the gutwall and liver. Accordingly, a suitable controlled release compositionmay be a composition that is designed to release tacrolimus in a delayedmanner so as to avoid or reduce the CYP3A4 metabolism in thegastrointestinal tract.

Delayed release is mainly brought about by some kind of enteric coating.Whereas semipermeable coating will show some kind of delayed release, itdoes not preciously enough “delay” release. Additionally it requires acertain amount of time to release the content. The coating sought forthis invention, is a pH dependant coating. This type of coating is veryresistant to release of drug until a certain pH is reached. Within veryfew 1/10′th of pH, the film alters properties and becomes permeable.Examples of pH-sensitive polymers, which are relatively insoluble andimpermeable at the pH of the stomach, but which are more soluble andpermeable at the pH of the small intestine and colon include, but notlimited to polyacrylamides, phthalate derivatives such as acidphthalates of carbohydrates, amylose acetate phthalate, celluloseacetate phthalate, other cellulose ester phthalates, cellulose etherphthalates, hydroxypropylcellulose phthalate,hydroxypropylethylcellulose phthalate, hydroxypropylmethylcellulosephthalate, methylcellulose phthalate, polyvinyl acetate phthalate,polyvinyl acetate hydrogen phthalate, sodium cellulose acetatephthalate, starch acid phthalate, styrene-maleic acid dibutyl phthalatecopolymer, styrene-maleic acid polyvinylacetate phthalate copolymer,styrene and maleic acid copolymers, polyacrylic acid derivatives such asacrylic acid and acrylic ester copolymers, polymethacrylic acid andesters thereof, poly acrylic methacrylic acid copolymers, shellac, andvinyl acetate and crotonic acid copolymers.

pH-sensitive polymers of specific interest include shellac; phthalatederivatives, particularly cellulose acetate phthalate, polyvinylacetatephthalate, and hydroxypropylmethylcellulose phthalate; polyacrylic acidderivatives, particularly polymethyl methacrylate blended with acrylicacid and acrylic ester copolymers; and vinyl acetate and crotonic acidcopolymers.

The release of the active substance from a composition having a delayedrelease coating could also be an enzymatic reaction, if for example Zeinor mono/di-glyceride mixtures are employed as coating material.

Upon oral administration to a mammal, including a human, in needthereof, a controlled release pharmaceutical composition according tothe present invention releases tacrolimus in such a manner that a plasmaconcentration of at least about 5 ng/ml such as, e.g., at least about7.5 ng/ml or at least about 10 ng/ml for a time period of at least about24 hours is obtained. In a specific aspect of the invention thedifference between the peak plasma concentration and plasmaconcentration measured 24 hours after administration is at the mostabout 20 ng/ml such as, e.g., at the most about 10 ng/ml, at the mostabout 7.5 ng/ml or at the most about 5 ng/ml.

In a specific aspect, the invention provides a pharmaceuticalcomposition or a solid dosage form that releases tacrolimus and/or ananalogue thereof relatively fast so as to enable a relatively fast onsetof therapeutic effect. In one aspect, the invention relates to apharmaceutical composition in particulate form comprising tacrolimusand/or an analogue thereof together with one or more pharmaceuticallyacceptable excipient, wherein the composition upon oral administrationto a mammal in need thereof in a controlled manner releases at leastabout 50% w/w of the total amount of tacrolimus or an analogue thereofwithin about 24 hours, such as, e.g., within about 22 hours, withinabout 20 hours, within about 18 hours, within about 15 hours or withinabout 12 hours.

In a further embodiment at the most about 60% w/w such as, e.g., at themost 62% w/w, at the most about 65% w/w or at the most about 70% w/wtacrolimus is released 15 hours after oral administration to a mammal ofa composition according to the invention or, alternatively, when testedin a suitable in vitro dissolution test, 15 hours after start of such atest.

More specifically, upon oral administration to a mammal in need thereofa composition according to the invention releases at least about 50% w/wof the total amount of tacrolimus and/or an analogue thereof withinabout 10 hours such as, e.g., within about 8 hours, within about 6hours, within about 4 hours or within about 3 hours.

In another embodiment, upon oral administration to a mammal in needthereof, a pharmaceutical composition according to the inventionreleases at least 80% w/w tacrolimus after about 0.5 hours or more suchas, e.g., after about 0.75 hours or more, about 1 hour or more, about 2hours or more, about 3 hours or more, about 4 hours or more or about 5hours or more; or alternatively, when tested in a suitable in vitrodissolution test releases at least 80% w/w after about 0.5 hours or moresuch as, e.g., after about 0.75 hours or more, about 1 hour or more,about 2 hours or more, about 3 hours or more, about 4 hours or more orabout 5 hours or more after start of the test.

In a further embodiment, upon oral administration to a mammal in needthereof a pharmaceutical composition according to the invention releasesat least about 55% w/w such as, e.g., about 60% w/w or more, about 65%w/w or more, about 70% w/w or more, about 75% w/w or more or about 80%w/w or more of the total amount of tacrolimus and/or an analogue thereofwithin about 24 hours, such as, e.g., within about 22 hours, withinabout 20 hours, within about 18 hours, about 15 hours, within about 12hours, within about 10 hours, within 8 hours or within about 6 hours.

Furthermore or alternatively, at least about 50% w/w of the total amountof tacrolimus and/or an analogue thereof is released about 24 hours,within about 22 hours, within about 20 hours, within about 18 hours,within 15 hours, within about 12 hours, when tested in an in vitrodissolution test and employing a dissolution medium comprising a bufferhaving pH 7.5. Guidance for a suitable dissolution test is described inthe Examples herein, but variations with respect to the specific methodemployed and the ingredients contained in the dissolution medium etc.are within the scope of the present invention. A person skilled in theart will know how to carry out a suitable dissolution test e.g. withguidance from USP, Ph. Eur. and the like. Suitable conditions for the invitro dissolution test are employing USP dissolution test (paddlemethod) and a buffer pH 7.5 containing 2.5% SDS and 1 g/mL of pancreatinas dissolution medium.

In other embodiments, the following conditions are fulfilled withrespect to in vitro dissolution test:

i) at least about 50% w/w of the total amount of tacrolimus or ananalogue thereof is released within about 10 hours such as, e.g., withinabout 8 hours, within about 6 hours, within about 4 hours, within about3 hours, within about 2 hours, within about 1 hour, within about 45 min,within about 30 min or within about 15 min, when tested in an in vitrodissolution test and employing a dissolution medium comprising a bufferhaving pH 7.5ii) at least about 50% w/w of the total amount of tacrolimus or ananalogue thereof is released within about 1.5 hours such as, e.g.,within about 1 hour, within about 0.75 hours, within about 0.5 hours orwithin about 20 minutes, when tested in an in vitro dissolution test andemploying a dissolution medium comprising a buffer having pH 7.5.iii) at least about 55% w/w such as, e.g., about 60% w/w or more, about65% w/w or more, about 70% w/w or more, about 75% w/w or more or about80% w/w or more of the total amount of tacrolimus or an analogue thereofis released within about 15 hours such as, e.g., within about 12 hours,within about 10 hours, within 8 hours or within about 6 hours, whentested in an in vitro dissolution test and employing a dissolutionmedium comprising a buffer having pH 7.5iv) at least about 55% w/w such as, e.g., about 60% w/w or more, about65% w/w or more, about 70% w/w or more, about 75% w/w or more or about80% w/w or more of the total amount of tacrolimus or an analogue thereofis released within about 5 hours such as, e.g., within about 4 hours,within about 3 hours, within about 2 hours, within about 1 hours orwithin about 30 minutes, when tested in an in vitro dissolution test andemploying a dissolution medium comprising a buffer having pH 7.5, and/orv) at least about 20% w/w such as, e.g., at least about 25% w/w, atleast about 30% w/w, at least about 35% w/w or at least about 40% w/w ofthe total amount of tacrolimus or an analogue thereof is released withinthe first 3 hours such as, e.g., within the first 2 hours or within thefirst hour when tested in an in vitro dissolution test and employing adissolution medium comprising a buffer having pH 7.5.

In an interesting embodiment, the composition is designed to have adelayed release of tacrolimus and/or an analogue thereof. Therefore, theinvention also includes a pharmaceutical composition in particulate formcomprising tacrolimus and/or an analogue thereof together with one ormore pharmaceutically acceptable excipient, wherein the composition uponoral administration to a mammal in need thereof has a delayed release oftacrolimus and/or an analogue thereof so that at the most 10% w/w suchas, e.g., at the most about 7.5% w/w or at the most about 5% w/w of thetotal amount of tacrolimus or an analogue thereof is released within thefirst two hours such as, e.g., within the first hour afteradministration.

In other embodiments, the following conditions are fulfilled withrespect to in vitro dissolution test performed under acidic conditions:

i) at the most about 30% w/w such as, e.g., at the most about 25% w/w,at the most about 20% w/w, at the most about 15% w/w or at the mostabout 10% w/w of tacrolimus or an analogue thereof is released within 2hours in an in vitro dissolution test employing a dissolution mediumhaving a pH of at the most about 5 such as, e.g. at the most about 4.5,at the most about 4, at the most about 3.5, at the most about 3, at themost about 2 or at the most about 1.5;ii) at the most about 10% w/w such as, e.g., at the most about 7.5% w/w,at the most about 5% w/w or at the most about 2.5% w/w of tacrolimus oran analogue thereof is released within 2 hours in an in vitrodissolution test employing a dissolution medium having a pH of at themost about 5 such as, e.g. at the most about 4.5, at the most about 4,at the most about 3.5, at the most about 3, at the most about 2 or atthe most about 1.5;iii) at the most about 60% w/w such as, e.g., at the most about 50% w/w,at the most about 40% w/w or at the most about 30% w/w of tacrolimus oran analogue thereof is released within 15 hours such as, e.g., withinabout 12 hours, when tested in an in vitro dissolution test employing adissolution medium having a pH of at the most about 4.5 such as, e.g. atthe most about 4.0, at the most about 3.5, at the most about 3, at themost about 2 or at the most about 1.5;iv) at the most about 40% w/w such as, e.g., at the most about 30% w/w,at the most about 25% w/w or at the most about 20% w/w of tacrolimus oran analogue thereof is released within 6 hours when tested in an invitro dissolution test employing a dissolution medium having a pH of atthe most about 4.5 such as, e.g. at the most about 4.0, at the mostabout 3.5, at the most about 3, at the most about 2 or at the most about1.5, and/orv) at the most about 30% w/w such as, e.g., at the most about 25% w/w,at the most about 20% w/w or at the most about 15% w/w of tacrolimus oran analogue thereof is released within 4 hours when tested in an invitro dissolution test employing a dissolution medium having a pH of atthe most about 4.5 such as, e.g. at the most about 4.0, at the mostabout 3.5, at the most about 3, at the most about 2 or at the most about1.5.

Apart from tacrolimus, a composition of the invention may also comprisea further therapeutically, prophylactically and/or diagnostically activesubstance. Notably combinations of tacrolimus with at least one of thefollowing active substances are of interest: Substances that areindicated for use in connection with organ transplantation such as,e.g., steroids, calcineurin inhibitors and/or anti-proliferative agents.Specific examples include prednisone, prednisolone, methylprednisone,cyclosporin, mycophenolate, azathioprine, sirolimus, everolimus,mycophenolate sodium, and FTY720 (Novartis).

The pharmaceutical compositions may be prepared by any convenient methodsuch as, e.g. granulation, mixing, spray drying etc. A particularlyuseful method is the method described in WO 03/004001. Herein isdescribed a process for the preparation of particulate material by acontrolled agglomeration method, i.e. a method, which enables acontrolled growth in particle size. The method involves spraying a firstcomposition comprising e.g. tacrolimus and a carrier, which has beenmelted, onto a second solid carrier medium. Normally, the meltablecarrier has a melting point of at least 5° C. but lower than the meltingpoint of tacrolimus. The melting point of the carrier may be in therange of 10° C. to 150° C., such as, e.g., in the range of 30° C. to100° C. or in the range of 40° C. to 50° C. is most preferred.

It is within the skills of the average practitioner to select a suitablecarrier being pharmaceutical acceptable, capable of dissolving or atleast partly dissolve tacrolimus and having a melting point in thedesired range using general knowledge and routine experimentation.Suitable candidate for carriers are described in WO 03/004001, which isherein incorporated by reference.

In the present context, suitable carriers are e.g. those mentioned as anoil or an oily-like material (as discussed later herein) as well asthose disclosed in WO 03/004001.

An advantage of using the controlled agglomeration method described inWO 03/004001 is that it is possible to apply a relatively large amountof a melt to a particulate material without having an undesirable growthin particle size. Accordingly, in one embodiment of the invention, theparticulate material of a pharmaceutical composition has a geometricweight mean diameter d_(gw) of ≧10 μm such as, e.g. ≧20 μm, from about20 to about 2000, from about 30 to about 2000, from about 50 to about2000, from about 60 to about 2000, from about 75 to about 2000 such as,e.g. from about 100 to about 1500 μm, from about 100 to about 1000 μm orfrom about 100 to about 700 μm, or at the most about 400 μm or at themost 300 μm such as, e.g., from about 50 to about 400 μm such as, e.g.,from about 50 to about 350 μm, from about 50 to about 300 μm, from about50 to about 250 μm or from about 100 to about 300 μm.

The particulate material obtained by the above-mentioned method hassuitable properties with respect to flowability and/or compressibilityand is therefore suitable for further processing into pharmaceuticaldosage forms.

Solid Dispersion and/or Solid Solution of Tacrolimus

The solid dispersion or solid dispersion used in a preferred embodimentof the invention comprises an active ingredient selected amongtacrolimus and analogues thereof, which ingredient is dispersed ordissolved in a hydrophilic or water-miscible vehicle having a meltingpoint (freezing point or pour point) of at least 20° C. in aconcentration of between about 0.01 w/w % and about 15 w/w %, and whichdispersion is forming a solid dispersion or solid solution at ambienttemperature (room temperature).

The concentration of the active ingredient in the hydrophilic orwater-miscible vehicle is at the most 15 w/w %, preferably at the most10 w/w %, preferably at the most 8 w/w %, more preferably at the most 6w/w %, even more preferably at the most 5 w/w %, at the most 4% w/w,especially at the most 3 w/w %, in particular at the most 2% w/w; and/oris at least about 0.05 w/w %, preferably at least about 0.1 w/w %, morepreferably at least about 0.5 w/w %, especially at least about 0.7 w/w%, in particular at least about 1 w/w %.

Physically, the combination of active ingredient and vehicle may eitherform a solid dispersion, i.e. the active ingredient is dispersed in thevehicle in particulate form, or may form a solid solution, i.e. theactive ingredient is dissolved in the vehicle at a molecular level. Theactive ingredient and the vehicle may also form a solid dispersionhaving therein a part of the active ingredient dissolved at a molecularlevel. The physical state of the dispersion and/or solution may bedetermined by using various techniques such as Hot Stage Microscopy(HSM), Differential Scanning calorimetry (DSC), Scanning ElectronMicroscopy (SEM) optionally in combination with Energy Dispersive X-ray(EDX), and X-ray powder diffraction. In a preferred embodiment, theactive ingredient is fully dissolved in the vehicle to form a solidsolution at ambient temperature.

The solid dispersion of the invention exhibits a very fast immediaterelease of tacrolimus, when a composition comprising the dispersion orsolution is tested in a dissolution test according to USP using anaqueous dissolution medium, and at least 50 w/w % of the activepharmaceutical ingredient is released within about 30 minutes,preferably within 20 minutes, more preferably within 15 minutes; such asat feast 75 w/w % of the active pharmaceutical ingredient is releasedwithin about 40 minutes, or even better at least 90 w/w % of the activepharmaceutical ingredient is released within about 60 minutes,preferably within 45 minutes. For example, the test may be carried outaccording to the any method and any specifications cited in USP. Thus,the dissolution test may be carried out in an aqueous dissolution mediumat a neutral or near-neutral pH, for example at pH 6.8, or at any acidicpH simulating the pH conditions in the gastrointestinal tract. However,variations with respect to the specific method employed and theingredients contained in the dissolution medium etc. are within thescope of the present invention. A person skilled in the art will knowhow to carry out a suitable dissolution test e.g. with guidance fromUSP, Ph. Eur. and the like. Suitable conditions for the in vitrodissolution test are employing USP dissolution test (paddle method) anda buffer pH 7.5 containing 2.5% SDS and 1 g/mL of pancreatin asdissolution medium.

The hydrophilic or water-miscible vehicle to be used according to theinvention is preferably one having a melting point (freezing point orpour point) of at least 20° C., more preferably at least 30° C., morepreferably at least 40° C., more preferably at least 50° C., even morepreferably at least 52° C., even more preferably at least 55° C., evenmore preferably at least 59° C., especially at least 61° C., inparticular at least 65° C.

Examples of useful hydrophilic or water-miscible vehicles to be usedaccording to this invention are selected from the group consisting ofpolyethylene glycols, polyoxyethylene oxides, poloxamers,polyoxyethylene stearates, poly-epsilon caprolactone, polyglycolizedglycerides such as Gelucire®, and mixtures thereof.

In a preferred embodiment of the invention, the vehicle is apolyethylene glycol (PEG), in particular a PEG having an averagemolecular weight of at least 1500, preferably at least 3000, morepreferably at least 4000, especially at least 6000. The polyethyleneglycol may advantageously be mixed with one or more other hydrophilic orwater-miscible vehicles, for example a poloxamer, preferably in aproportion (on a weight/weight basis) of between 1:3 and 10:1,preferably between 1:1 and 5:1, more preferably between and 3:2 4:1,especially between 2:1 and 3:1, in particular about 7:3. A specificexample of a useful mixture is a mixture of PEG6000 and poloxamer 188 inthe ratio 7:3.

For polyethylene glycols (PEG), the melting point (freezing point orpour point) increases as the average molecular weight increases. Forexample, PEG 400 is in the range of 4-8° C., PEG 600 is in the range of20-25° C., PEG1500 is in the range of 44-48° C., PEG2000 is about 52°C., PEG 4000 is about 59° C., PEG 6000 is about 65° C. ad PEG 8000 isabout 61° C.

Useful poloxamers (also denoted polyoxypropylene-polyoxyethylene blockcopolymers) are for example poloxamer 188, poloxamer 237, poloxamer 338or poloxamer 407 or other block copolymers of ethylene oxide andpropylene oxide such as the Pluronic® and/or Tetronic® series. Suitableblock copolymers of the Pluronic® series include polymers having amolecular weight of about 3,000 or more such as, e.g. from about 4,000to about 20,000 and/or a viscosity (Brookfield) from about 200 to about4,000 cps such as, e.g., from about 250 to about 3,000 cps. Suitableexamples include Pluronic® F38, P65, P68LF, P75, F77, P84, P85, F87,F88, F98, P103, P104, P105, F108, P123, F123, F127, 10R8, 17R8, 25R5,25R8 etc. Suitable block copolymers of the Tetronic® series includepolymers having a molecular weight of about 8,000 or more such as, e.g.,from about 9,000 to about 35,000 and/or a viscosity (Brookfield) of fromabout 500 to about 45,000 cps such as, e.g., from about 600 to about40,000. The viscosities given above are determined at 60° C. forsubstances that are pastes at room temperature and at 77° C. forsubstances that are solids at room temperature.

In a preferred embodiment of the present invention, the poloxamer ispoloxamer 188, which has an average molecular weight of about 8400 and amelting point of about 50-54° C.

Other useful hydrophilic or water-miscible vehicles may bepolyvinylpyrrolidones, polyvinyl-polyvinylacetate copolymers (PVP-PVA),polyvinyl alcohol (PVA), polymethacrylic polymers (Eudragit RS; EudragitRL, Eudragit NE, Eudragit E), cellulose derivatives includinghydroxypropyl methylcellulose (HPMC), hydroxypropyl cellulose (HPC),methylcellulose, sodium carboxymethylcellulose, hydroxyethyl cellulose,pectins, cyclodextrins, galactomannans, alginates, carragenates, xanthangums and mixtures thereof.

“Polyglycolized glycerides” denotes a mixture of mono-, di- andtriglycerides and polyethylene glycol (PEG) mono- and diesters,preferably of molecular weight between 200 and 600, where appropriate offree glycerol and free PEG, whose HLB value is adjusted by the length ofthe PEG chain, and whose melting point is adjusted by the length of thechains of the fatty acids, of the PEG and by the degree of saturation ofthe fatty chains, and hence of the starting oil; examples of suchmixtures are Gelucire®. Gelucire®compositions are inert semi-solid waxymaterials which are amphiphilic in character and are available withvarying physical characteristics. They are surface active in nature anddisperse or solubilize in aqueous media forming micelles, microscopicglobules or vesicles. They are identified by their melting point/HLBvalue. The melting point is expressed in degrees Celsius and the HLB(Hydrophile-Lipophile Balance) is a numerical scale extending from 0 toapproximately 20. Lower HLB values denote more lipophilic andhydrophobic substances, and higher values denote more hydrophilic andlipophobic substances. The affinity of a compound for water or for oilysubstances is determined and its HLB value is assigned experimentally.One or a mixture of different grades of Gelucire® excipient may bechosen to achieve the desired characteristics of melting point and/orHLB value. They are mixtures of monoesters, diesters and/or triesters ofglycerides of long chain (C₁₂ to C₁₈) fatty acids, and PEG (mono- and/ordi) esters of long chain (C₁₂ to C₁₈) fatty acids and can include freePEG. Gelucire® compositions are generally described as fatty acid estersof glycerol and PEG esters or as polyglycolized glycerides. Gelucire®compositions are characterized by a wide range of melting points of fromabout 33° C. to about 64° C. and most commonly from about 35° C. toabout 55° C., and by a variety of HLB values of from about 1 to about14, most commonly from about 7 to about 14. For example, Gelucire® 50/13designates a melting point of approximately 50° C. and an HLB value ofabout 13 to this grade of Gelucire®.

Pharmaceutically Acceptable Excipients

Examples of suitable excipients for use in a composition or solid dosageform according to the present invention include fillers, diluents,disintegrants, binders, lubricants and the like and mixtures thereof. Asthe composition or solid dosage form according to the invention may beused for different purposes, the choice of excipients is normally madetaken such different uses into considerations. Other pharmaceuticallyacceptable excipients for suitable use are e.g. acidifying agents,alkalizing agents, preservatives, antioxidants, buffering agents,chelating agents, coloring agents, complexing agents, emulsifying and/orsolubilizing agents, flavors and perfumes, humectants, sweeteningagents, wetting agents and the like.

Examples of suitable fillers, diluents and/or binders include lactose(e.g. spray-dried lactose, α-lactose, β-lactose, Tabletose®, variousgrades of Pharmatose®, Microtose® or Fast-Floc®), microcrystallinecellulose (various grades of Avicel®, Elcema®, Vivacel®, Ming Tai® orSolka-Floc®), hydroxypropylcellulose, L-hydroxypropylcellulose (lowsubstituted), hydroxypropyl methylcellulose (HPMC) (e.g. Methocel E, Fand K, Metolose SH of Shin-Etsu, Ltd, such as, e.g. the 4,000 cps gradesof Methocel E and Metolose 60 SH, the 4,000 cps grades of Methocel F andMetolose 65 SH, the 4,000, 15,000 and 100,000 cps grades of Methocel K;and the 4,000, 15,000, 39,000 and 100,000 grades of Metolose 90 SH),methylcellulose polymers (such as, e.g., Methocel A, Methocel A4C,Methocel A15C, Methocel A4M), hydroxyethylcellulose, sodiumcarboxymethylcellulose, carboxymethylene,carboxymethylhydroxyethylcellulose and other cellulose derivatives,sucrose, agarose, sorbitol, mannitol, dextrins, maltodextrins, starchesor modified starches (including potato starch, maize starch and ricestarch), calcium phosphate (e.g. basic calcium phosphate, calciumhydrogen phosphate, dicalcium phosphate hydrate), calcium sulfate,calcium carbonate, sodium alginate, collagen etc.

Specific examples of diluents are e.g. calcium carbonate, dibasiccalcium phosphate, tribasic calcium phosphate, calcium sulfate,microcrystalline cellulose, powdered cellulose, dextrans, dextrin,dextrose, fructose, kaolin, lactose, mannitol, sorbitol, starch,pregelatinized starch, sucrose, sugar etc.

Specific examples of disintegrants are e.g. alginic acid or alginates,microcrystalline cellulose, hydroxypropyl cellulose and other cellulosederivatives, croscarmellose sodium, crospovidone, polacrillin potassium,sodium starch glycolate, starch, pregelatinized starch, carboxymethylstarch (e.g. Primogel® and Explotab®) etc.

Specific examples of binders are e.g. acacia, alginic acid, agar,calcium carrageenan, sodium carboxymethylcellulose, microcrystallinecellulose, dextrin, ethylcellulose, gelatin, liquid glucose, guar gum,hydroxypropyl methylcellulose, methylcellulose, pectin, PEG, povidone,pregelatinized starch etc.

Glidants and lubricants may also be included in the composition.Examples include stearic acid, magnesium stearate, calcium stearate orother metallic stearate, talc, waxes and glycerides, light mineral oil,PEG, glyceryl behenate, colloidal silica, hydrogenated vegetable oils,corn starch, sodium stearyl fumarate, polyethylene glycols, alkylsulfates, sodium benzoate, sodium acetate etc.

Other excipients which may be included in a composition or solid dosageform of the invention are e.g. flavoring agents, coloring agents,taste-masking agents, pH-adjusting agents, buffering agents,preservatives, stabilizing agents, anti-oxidants, wetting agents,humidity-adjusting agents, surface-active agents, suspending agents,absorption enhancing agents, agents for modified release etc.

Other additives in a composition or a solid dosage form according to theinvention may be antioxidants like e.g. ascorbic acid, ascorbylpalmitate, butylated hydroxyanisole, butylated hydroxytoluene,hypophosphorous acid, monothioglycerol, potassium metabisulfite, propylgallate, sodium formaldehyde sulfoxylate, sodium metabisulfite, sodiumthiosulfate, sulfur dioxide, tocopherol, tocopherol acetate, tocopherolhemisuccinate, TPGS or other tocopherol derivatives, etc. The carriercomposition may also contain e.g. stabilising agents. The concentrationof an antioxidant and/or a stabilizing agent in the carrier compositionis normally from about 0.1% w/w to about 5% w/w.

A composition or solid dosage form according to the invention may alsoinclude one or more surfactants or substances having surface-activeproperties. It is contemplated that such substances are involved in thewetting of the slightly soluble active substance and thus, contributesto improved solubility characteristics of the active substance.

Suitable excipients for use in a composition or a solid dosage formaccording to the invention are surfactants such as, e.g., amphiphilicsurfactants as those disclosed in WO 00/50007 in the name of Lipocine,Inc. Examples of suitable surfactants are

-   -   i) polyethoxylated fatty acids such as, e.g. fatty acid mono- or        diesters of polyethylene glycol or mixtures thereof such as,        e.g. mono- or diesters of polyethylene glycol with lauric acid,        oleic acid, stearic acid, myristic acid, ricinoleic acid, and        the polyethylene glycol may be selected from PEG 4, PEG 5, PEG        6, PEG 7, PEG 8, PEG 9, PEG 10, PEG 12, PEG 15, PEG 20, PEG 25,        PEG 30, PEG 32, PEG 40, PEG 45, PEG 50, PEG 55, PEG 100, PEG        200, PEG 400, PEG 600, PEG 800, PEG 1000, PEG 2000, PEG 3000,        PEG 4000, PEG 5000, PEG 6000, PEG 7000, PEG 8000, PEG 9000, PEG        1000, PEG 10,000, PEG 15,000, PEG 20,000, PEG 35,000,    -   ii) polyethylene glycol glycerol fatty acid esters, i.e. esters        like the above-mentioned but in the form of glyceryl esters of        the individual fatty acids;    -   iii) glycerol, propylene glycol, ethylene glycol, PEG or        sorbitol esters with e.g. vegetable oils like e.g. hydrogenated        castor oil, almond oil, palm kernel oil, castor oil, apricot        kernel oil, olive oil, peanut oil, hydrogenated palm kernel oil        and the like,    -   iv) polyglycerized fatty acids like e.g. polyglycerol stearate,        polyglycerol oleate, polyglycerol ricinoleate, polyglycerol        linoleate,    -   v) propylene glycol fatty acid esters such as, e.g. propylene        glycol monolaurate, propylene glycol ricinoleate and the like,    -   vi) mono- and diglycerides like e.g. glyceryl monooleate,        glyceryl dioleae, glyceryl mono- and/or dioleate, glyceryl        caprylate, glyceryl caprate etc.;    -   vii) sterol and sterol derivatives;    -   viii) polyethylene glycol sorbitan fatty acid esters        (PEG-sorbitan fatty acid esters) such as esters of PEG with the        various molecular weights indicated above, and the various        Tween® series;    -   ix) polyethylene glycol alkyl ethers such as, e.g. PEG oleyl        ether and PEG lauryl ether;    -   x) sugar esters like e.g. sucrose monopalmitate and sucrose        monolaurate;    -   xi) polyethylene glycol alkyl phenols like e.g. the Triton® X or        N series;    -   xii) polyoxyethylene-polyoxypropylene block copolymers such as,        e.g., the Pluronic® series, the Synperonic® series, Emkalyx®,        Lutrol®, Supronic® etc. The generic term for these polymers is        “poloxamers” and relevant examples in the present context are        Poloxamer 105, 108, 122, 123, 124, 181, 182, 183, 184, 185, 188,        212, 215, 217, 231, 234, 235, 237, 238, 282, 284, 288, 331, 333,        334, 335, 338, 401, 402, 403 and 407;    -   xiii) sorbitan fatty acid esters like the Span® series or        Ariacel® series such as, e.g. sorbinan monolaurate, sorbitan        monopalmitate, sorbitan monooleate, sorbitan monostearate etc.;    -   xiv) lower alcohol fatty acid esters like e.g. oleate, isopropyl        myristate, isopropyl palmitate etc.;    -   xv) ionic surfactants including cationic, anionic and        zwitterionic surfactants such as, e.g. fatty acid salts, bile        salts, phospholipids, phosphoric acid esters, carboxylates,        sulfates and sulfonates etc.

When a surfactant or a mixture of surfactants is present in acomposition or a solid dosage form of the invention, the concentrationof the surfactant(s) is normally in a range of from about 0.1-80% w/wsuch as, e.g., from about 0.1 to about 20% w/w, from about 0.1 to about15% w/w, from about 0.5 to about 10% w/w, or alternatively, from about0.10 to about 80% w/w such as, e.g. from about 10 to about 70% w/w, fromabout 20 to about 60% w/w or from about 30 to about 50% w/w.

In a specific aspect of the invention, the at least one of the one ormore pharmaceutically acceptable excipient is selected from the groupconsisting of silica acid or a derivative or salt thereof includingsilicates, silicon dioxide and polymers thereof; magnesiumaluminosilicate and/or magnesium aluminometasilicate, bentonite, kaolin,magnesium trisilicate, montmorillonite and/or saponite.

Such materials are is especially useful as a sorption material for oilsor oily-like materials in pharmaceuticals, cosmetics and/or foodstuff.In a specific embodiment, the material is used as a sorption materialfor oils or oily-like materials in pharmaceuticals. The material thathas the ability to function as a sorption material for oils or oily-likematerials is also denoted “oil sorption material”. Furthermore, in thepresent context the term “sorption” is used to denote “absorption” aswell as “adsorption”. It should be understood that whenever one of theterms is used it is intended to cover the phenomenon absorption as wellas adsorption.

Notably, the pharmaceutically acceptable excipient may comprise a silicaacid or a derivative or salt thereof such as, e.g., silicon dioxide or apolymer thereof as a pharmaceutically acceptable excipient. Dependent onthe quality employed a silicon dioxide may be a lubricant or it may bean oil sorption material. Qualities fulfilling the latter function seemto be most important.

In a specific embodiment, a composition or solid dosage form accordingto invention comprises a pharmaceutically acceptable excipient that is asilicon dioxide product that has properties corresponding to Aeroperl®300 (available from Degussa, Frankfurt, Germany). As it appears from theexamples herein, a very suitable material is Aeroperl® 300 (includingmaterials with properties like or corresponding to those of Aeroperl®300).

Use of an oil sorption material in compositions or dosage formsaccording to the invention is very advantageous for the preparation ofpharmaceutical, cosmetic, nutritional and/or food compositions, whereinthe composition comprises oil or an oily-like material. One of theadvantages is that is it possible to incorporate a relatively largeamount of oil and oily-like material and still have a material that issolid. Thus, it is possible to prepare solid compositions with arelatively high load of oil or oily-like materials by use of an oilsorption material according to the invention. Within the pharmaceuticalfield it is an advantage to be able to incorporate a relatively largeamount of an oil or an oily-like material in a solid compositionespecially in those situation where the active substance does not havesuitable properties with respect to water solubility (e.g. poor watersolubility), stability in aqueous medium (i.e. degradation occurs inaqueous medium), oral bioavailability (e.g. low bioavailability) etc.,or in those situations where it is desired to modify the release of anactive substance from a composition in order to obtain a controlled,delayed, sustained and/or pulsed delivery of the active substance. Thus,in a specific embodiment it is used in the preparation of pharmaceuticalcompositions.

The oil sorption material for use in the processing into solidcompositions normally absorbs about 5% w/w or more, such as, e.g., about10% w/w or more, about 15% w/w or more, about 20% w/w or more, about 25%w/w or more, about 30% w/w or more, about 35% w/w or more, about 40% w/wor more, about 45% w/w or more, about 50 w/w or more, about 55% w/w ormore, about 60% w/w or more, about 65% w/w or more, about 70% w/w ormore, about 75% w/w or more, about 80% w/w or more, about 85% w/w ormore, about 90% w/w or more or about 95% w/w or more of an oil or anoily material and is still a solid material.

Yet another aspect of the invention relates to compositions or soliddosage forms comprising an oil or an oily material.

In the present context the term “oils and oily materials” is used in avery broad sense including oils, waxes, semi-solid materials andmaterials that normally are used as solvents (such as organic solvents)or cosolvents within the pharmaceutical industry, and the term alsoincludes therapeutically and/or prophylactically active substances thatare in liquid form at ambient temperature; furthermore the term includesemulsions like e.g. microemulsions and nanoemulsions and suspensions.The oils and oily-like materials that can be absorbed will normally beliquid at ambient or elevated temperature (for practical reasons themax. temperature is about 250° C.). They may be hydrophilic, lipophilic,hydrophobic and/or amphiphilic materials.

The oils and oily-like material that are suitable for use in the presentcontext are substances or materials, which have a melting point of atleast about 0° C. and at the most about 250° C.

In specific embodiments of the invention, the oil or oily-like materialhas a melting point of about 5° C. or more such as, e.g., about 10° C.or more, about 15° C. or more, about 20° C. or more or about 25° C. ormore.

In further embodiments of the invention, the oil or oily-like materialhas a melting point of at least about 25° C. such as, e.g., at leastabout 30° C. at least about 35° C. or at least about 40° C. Forpractical reasons, the melting point may normally not be too high, thus,the oil or oily-like material normally has a melting point of at themost about 300° C. such as, e.g., at the most about 250° C., at the mostabout 200° C., at the most about 150° C. or at the most about 100° C. Ifthe melting point is higher a relatively high temperature may promotee.g. oxidation or other kind of degradation of an active substance inthose cases where e.g. a therapeutically and/or prophylactically activesubstance is included.

In the present context, the melting point is determined by DSC(Differential Scanning calorimetry). The melting point is determined asthe temperature at which the linear increase of the DSC curve intersectsthe temperature axis.

Interesting oils or oily-like materials are generally substances, whichare used in the manufacture of pharmaceuticals as so-called melt bindersor solid solvents (in the form of solid dosage form), or as co-solventsor ingredients in pharmaceuticals for topical use.

It may be hydrophilic, hydrophobic and/or have surface-activeproperties. In general hydrophilic and/or hydrophobic oils or oily-likematerials are suitable for use in the manufacture of a pharmaceuticalcomposition comprising a therapeutically and/or prophylactically activesubstance that has a relatively low aqueous solubility and/or when therelease of the active substance from the pharmaceutical composition isdesigned to be immediate or non-modified. Hydrophobic oil or oily-likematerials, on the other hand, are normally used in the manufacture of amodified release pharmaceutical composition. The above-givenconsiderations are simplified to illustrate general principles, butthere are many cases where other combinations of oils or oily-likematerials and other purposes are relevant and, therefore, the examplesabove should not in any way limit the invention.

Typically, a suitable hydrophilic oil or oily-like material is selectedfrom the group consisting of: polyether glycols such as, e.g.,polyethylene glycols, polypropylene glycols; polyoxyethylenes;polyoxypropylenes; poloxamers and mixtures thereof, or it may beselected from the group consisting of: xylitol, sorbitol, potassiumsodium tartrate, sucrose tribehenate, glucose, rhamnose, lactitol,behenic acid, hydroquinon monomethyl ether, sodium acetate, ethylfumarate, myristic acid, citric acid, Gelucire 50/13, other Geluciretypes such as, e.g., Gelucire 44/14 etc., Gelucire 50/10, Gelucire62/05, Sucro-ester 7, Sucro-ester 11, Sucro-ester 15, maltose, mannitoland mixtures thereof.

A suitable hydrophobic oil or oily-like material may be selected fromthe group consisting of: straight chain saturated hydrocarbons, sorbitanesters, paraffins; fats and oils such as e.g., cacao butter, beeftallow, lard, polyether glycol esters; higher fatty acid such as, e.g.stearic acid, myristic acid, palmitic acid, higher alcohols such as,e.g., cetanol, stearyl alcohol, low melting point waxes such as, e.g.,glyceryl monostearate, glyceryl monooleate, hydrogenated tallow,myristyl alcohol, stearyl alcohol, substituted and/or unsubstitutedmonoglycerides, substituted and/or unsubstituted diglycerides,substituted and/or unsubstituted triglycerides, yellow beeswax, whitebeeswax, carnauba wax, castor wax, japan wax, acetylate monoglycerides;NVP polymers, PVP polymers, acrylic polymers, or a mixture thereof.

In an interesting embodiment, the oil or oily-like material is apolyethylene glycol having an average molecular weight in a range offrom about 400 to about 35,000 such as, e.g., from about 800 to about35,000, from about 1,000 to about 35,000 such as, e.g., polyethyleneglycol 1,000, polyethylene glycol 2,000, polyethylene glycol 3,000,polyethylene glycol 4,000, polyethylene glycol 5,000, polyethyleneglycol 6000, polyethylene glycol 7,000, polyethylene glycol 8,000,polyethylene glycol 9,000 polyethylene glycol 10,000, polyethyleneglycol 15,000, polyethylene glycol 20,000, or polyethylene glycol35,000. In certain situations polyethylene glycol may be employed with amolecular weight from about 35,000 to about 100,000.

In another interesting embodiment, the oil or oily-like material ispolyethylene oxide having a molecular weight of from about 2,000 toabout 7,000,000 such as, e.g. from about 2,000 to about 100,000, fromabout 5,000 to about 75,000, from about 10,000 to about 60,000, fromabout 15,000 to about 50,000, from about 20,000 to about 40,000, fromabout 100,000 to about 7,000,000 such as, e.g., from about 100,000 toabout 1,000,000, from about 100,000 to about 600,000, from about 100,000to about 400,000 or from about 100,000 to about 300,000.

In another embodiment, the oil or oily-like material is a poloxamer suchas, e.g. Poloxamer 188, Poloxamer 237, Poloxamer 338 or Poloxamer 407 orother block copolymers of ethylene oxide and propylene oxide such as thePluronic® and/or Tetronic® series. Suitable block copolymers of thePluronic® series include polymers having a molecular weight of about3,000 or more such as, e.g. from about 4,000 to about 20,000 and/or aviscosity (Brookfield) from about 200 to about 4,000 cps such as, e.g.,from about 250 to about 3,000 cps. Suitable examples include Pluronic®F38, P65, P68LF, P75, F77, P84, P85, F87, F88, F98, P103, P104, P105,F108, P123, F123, F127, 10R8, 17R8, 25R5, 25R8 etc. Suitable blockcopolymers of the Tetronic® series include polymers having a molecularweight of about 8,000 or more such as, e.g., from about 9,000 to about35,000 and/or a viscosity (Brookfield) of from about 500 to about 45,000cps such as, e.g., from about 600 to about 40,000. The viscosities givenabove are determined at 60° C. for substances that are pastes at roomtemperature and at 77° C. for substances that are solids at roomtemperature.

The oil or oily-like material may also be a sorbitan ester such as,e.g., sorbitan di-isostearate, sorbitan dioleate, sorbitan monolaurate,sorbitan monoisostearate, sorbitan monooleate, sorbitan monopalmitate,sorbitan monostearate, sorbitan sesqui-isostearate, sorbitansesquioleate, sorbitan sesquistearate, sorbitan tri-isostearate,sorbitan trioleate, sorbitan tristearate or mixtures thereof.

The oil or oily-like material may of course comprise a mixture ofdifferent oils or oily-like materials such as, e.g., a mixture ofhydrophilic and/or hydrophobic materials.

Other suitable oils or oily-like materials may be solvents or semi-solidexcipients like, e.g. propylene glycol, polyglycolised glyceridesincluding Gelucire 44/14, complex fatty materials of plant originincluding theobroma oil, carnauba wax, vegetable oils like e.g. almondoil, coconut oil, corn oil, cottonseed oil, sesame oil, soya oil, oliveoil, castor oil, palm kernels oil, peanut oil, rape oil, grape seed oiletc., hydrogenated vegetable oils such as, e.g. hydrogenated peanut oil,hydrogenated palm kernels oil, hydrogenated cottonseed oil, hydrogenatedsoya oil, hydrogenated castor oil, hydrogenated coconut oil; naturalfatty materials of animal origin including beeswax, lanolin, fattyalcohols including cetyl, stearyl, lauric, myristic, palmitic, stearicfatty alcohols; esters including glycerol stearate, glycol stearate,ethyl oleate, isopropyl myristate; liquid interesterified semi-syntheticglycerides including Miglycol 810/812; amide or fatty acid alcolamidesincluding stearamide ethanol, diethanolamide of fatty coconut acids,acetic acid esters of mono and di-glycerides, citric acid esters of monoand di-glycerides, lactic acid esters of mono and diglycerides, mono anddi-glycerides, poly-glycerol esters of fatty acids, poly-glycerolpoly-ricinoleate, propylene glycol esters of fatty acids, sorbitanmonostearates, sorbitan tristearates, sodium stearoyl lactylates,calcium stearoyl lactylates, diacetyl tartaric acid esters of mono anddi-glycerides etc.

Normally, a pharmaceutical composition or a solid dosage form accordingto the invention has a concentration of the oil or oily-like material inthe composition of about 5% w/w or more such as, e.g., about 10% w/w ormore, about 15% w/w or more, about 20% w/w or more, about 25% w/w ormore, about 30% w/w or more, about 35% w/w or more, about 40% w/w ormore, about 45% w/w or more, about 50 w/w or more, about 55% w/w ormore, about 60% w/w or more, about 65% w/w or more, about 70% w/w ormore, about 75% w/w or more, about 80% w/w or more, about 85% w/w ormore, about 90% w/w or more or about 95% w/w or more.

In specific embodiments the concentration of the oil or oily-likematerial in a composition or solid dosage form of the invention is in arange from about 20% to about 80% w/w such as, e.g., from about 25% toabout 75% w/w.

One of the advantages is that is it possible to incorporate a relativelylarge amount of oil and oily-like material and still have a materialthat is solid. Thus, it is possible to prepare solid compositions with arelatively high load of oil or oily-like materials by use of an oilsorption material according to the invention. Within the pharmaceuticalfield it is an advantage to be able to incorporate a relatively largeamount of an oil or an oily-like material in a solid compositionespecially in those situation where the active substance does not havesuitable properties with respect to water solubility (e.g. poor watersolubility), stability in aqueous medium (i.e. degradation occurs inaqueous medium), oral bioavailability (e.g. low bioavailability) etc.,or in those situations where it is desired to modify the release of anactive substance from a composition in order to obtain a controlled,delayed, sustained and/or pulsed delivery of the active substance.

A further advantage is that the particulate material obtained is afree-flowing powder and therefore readily processable into e.g. soliddosage forms such as tablets, capsules or sachets. Normally, theparticulate material has properties that are suitable in order tomanufacture tablets by direct compression without addition of largeamounts of further additives. A suitable test for testing theflowability of the particulate material is the method described in Ph.Eur. and measuring the flow rate of the material out of a funnel with anozzle (orifice) diameter of 10.0 mm.

In an important embodiment of the invention, at least a part oftacrolimus and/or an analogue thereof is present in the composition inthe form of a solid solution including a molecular dispersion and asolid dispersion. Normally, 10% or more such as, e.g., 20% or more, 30%or more, 40% or more, 50% or more, 60% or more, 70% or more, 80% ormore, 90% or more such as, e.g., 95% or more or about 100% w/w oftacrolimus and/or an analogue thereof is present in the composition inthe form of a solid dispersion.

A solid dispersion may be obtained in different ways e.g. by employingorganic solvents or by dispersing or dissolving the active substance inanother suitable medium (e.g. an oil or an oily-like material that is inliquid form at room temperature or at elevated temperatures).

Solid dispersions (solvent method) may for example be prepared bydissolving a physical mixture of the active substance (e.g. a drugsubstance) and the carrier in a common organic solvent, followed byevaporation of the solvent. The carrier is often a hydrophilic polymer.Suitable organic solvents include pharmaceutical acceptable solvent inwhich the active substance is soluble such as methanol, ethanol,methylene chloride, chloroform, ethylacetate, acetone or mixturesthereof.

Suitable water soluble carriers include polymers such as polyethyleneglycol, poloxamers, polyoxyethylene stearates, poly-ε-caprolactone,polyvinylpyrrolidone (PVP), polyvinylpyrrolidone-polyvinylacetatecopolymer PVP-PVA (Kollidon VA64), poly-methacrylic polymers (EudragitRS, Eudragit RL, Eudragit NE, Eudragit E) and polyvinyl alcohol (PVA),hydroxypropyl cellulose (HPC), hydroxypropyl methyl cellulose (HPMC),methyl cellulose, and poly(ethylene oxide) (PEO).

Polymers containing acidic functional groups may be suitable for soliddispersions, which release the active substance in a preferred pH rangeproviding acceptable absorption in the intestines. Such polymers may beone or more selected from the group comprising hydroxypropylmethylcellulose phthalate (HMPCP), polyvinyl acetate phthalate (PVAP),hydroxypropylmethylcellulose acetate succinate (HPMCAS), alginate,carbomer, carboxymethylcellulose, methacrylic acid copolymer (EudragitL, Eudragit S), shellac, cellulose acetate phthalate (CAP), starchglycolate, polacrylin, methyl cellulose acetate phthalate,hydroxypropylcellulose acetate phthalate, cellulose acetateterephthalate, cellulose acetate isophthalate and cellulose acetatetrimellitate.

Relative to the amount of the active substance and the polymer in thesolid dispersion, the weight ratio of active substance to polymer may bein a range of from about 3:1 to about 1:20. However, narrower ranger offrom about 3:1 to about 1:5, such as, e.g., from about 1:1 to about 1:3or about may also be used.

The solid dispersion is preferably formed by spray drying techniques,controlled agglomeration, freeze-drying or coating on carrier particlesor any other solvent removal process. The dried product contains theactive substance present in the form of a solid dispersion including amolecular dispersion and a solid solution.

As an alternative to the use of organic solvents the drug and polymermay be co-grinded or extruded at elevated temperatures (melt extrusion).

The pharmaceutical compositions comprising tacrolimus at least partly inform of a solid dispersion or solution may in principle be preparedusing any suitable procedure for preparing pharmaceutical compositionsknown within the art.

Apart from using the organic solvent based method, solid dispersion orsolid solutions of tacrolimus and/or an analogue thereof may be obtainedby dispersing and/or dissolving tacrolimus in the carrier compositionused in the controlled agglomeration method. Stabilizing agents etc. maybe added in order to ensure the stability of the soliddispersion/solution.

In another aspect, the invention relates to a method for the preparationof a pharmaceutical composition according to the invention. In general,any suitable method within the pharmaceutical field may be employed.However, in order to enable incorporation of a relatively high amount ofan oil or an oily-like material especially the method described in WO03/004001 has proved useful. WO 03/004001 is hereby incorporated byreference. The method comprises spraying a first composition in liquidform, said composition comprising a first vehicle or carrier and havinga melting point above 5° C. onto a second composition comprising asecond support or carrier material, said second composition e.g. beingin the fluidised state and having a temperature below the melting pointof the first vehicle or carrier. The active substance may be present inthe first vehicle or carrier composition and/or in the second support orcarrier composition. However, in those cases where tacrolimus and/or ananalogue thereof are present, at least partly, in the form of as a soliddispersion, it is advantageous to incorporate or dissolve tacrolimusand/or an analogue thereof in the first vehicle or carrier composition.

Solid Dosage Forms

The pharmaceutical composition according to the invention is inparticulate form and may be employed as such. However, in many cases itis more convenient to present the composition in the form of granules,pellets, microspheres, nanoparticles and the like or in the form ofsolid dosage forms including tablets, capsules and sachets and the like.A solid dosage form according to the invention may be a single unitdosage form or it may in the form of a polydepot dosage form contain amultiplicity of individual units such as, e.g., pellets, beads and/orgranules.

Normally, a pharmaceutical composition or a solid dosage form of theinvention is intended for administration via the oral, buccal orsublingual administration route.

The invention also relates to the above-mentioned presentation form.Within the scope of the invention are compositions/solid dosage formsthat are intended to release tacrolimus and/or an analogue thereof in afast release, a delayed release or modified release manner. All of thesemanners are considered to be a controlled manner. Further, a pHdependants release is also covered by the term “controlled manner”.

A solid dosage form according to the present invention comprises apharmaceutical composition in particulate form as described above. Thedetails and particulars disclosed under this main aspect of theinvention apply mutatis mutandis to the other aspects of the invention.Accordingly, the properties with respect to increase in bioavailability,changes in bioavailability parameters, reduction in adverse food effectas well as release of tacrolimus and/or an analogue thereof etc.described and/or claimed herein for pharmaceutical compositions inparticulate form are analogues for a solid dosage form according to thepresent invention.

Normally, the concentration of the pharmaceutical composition inparticulate form is in a range of from about 5 to 100% w/w such as,e.g., from about 10% to about 90% w/w, from about 15% to about 85% w/w,from about 20% to about 80% w/w, from about 25% to about 80% w/w, fromabout 30% to about 80% w/w, from about 35% to about 80% w/w, from about40% to about 75% w/w, from about 45% to about 75% w/w or from about 50%to about 70% w/w of the dosage form. In an embodiment of the invention,the concentration of the pharmaceutical composition in particulate formis 50% w/w or more of the dosage form.

A solid dosage form according to the invention is obtained by processingthe particulate material according to the invention by means oftechniques well-known to a person skilled in the art. Normally, itinvolves further addition of one or more of the pharmaceuticallyacceptable excipients mentioned herein.

The composition or solid dosage form according to the invention may bedesigned to release tacrolimus and/or an analogue thereof in anysuitable manner provided that the increase in bioavailability ispresent. Thus, the active substance may be released relatively fast inorder to obtain an enhanced on-set of action, it may be released so asto follow zero or first order kinetics or it may be released in amodified manner in order to obtain a predetermined pattern of release.All of these ways are considered controlled manners. Plain formulationsare also within the scope of the present invention.

The recommended dosage range for Prograf® is 0.1 to 0.2 mg/kg/day givenevery 12 hours in two divided doses. More importantly the blood levelshas to be monitored.

The typical level for 1-3 months is 7-0.20 ng/mL and 4-12 months thelevels should be 5-15 ng/mL. This is only guiding values and may varyfrom types of transplant and ethnicity.

The following was found for kidney transplant patients.

Caucasian n = 114 Black n = 56 Trough Trough Time After DoseConcentrations Dose Concentrations Transplant (mg/kg) (ng/mL) (mg/kg)(ng/mL) Day 7 0.18 12.0 0.23 10.9 Month 1 0.17 12.8 0.26 12.9 Month 60.14 11.8 0.24 11.5 Month 12 0.13 10.1 0.19 11.0

The expected dosage recommendation for products of the present inventionwill be from 0.02 mg/kg/day to 0.15 mg/kg/day, dosed once a day.

The composition or solid dosage form according to the invention may alsobe coated with a film coating, an enteric coating, a modified releasecoating, a protective coating, an anti-adhesive coating etc.

A solid dosage form according to the invention may also be coated inorder to obtain suitable properties e.g. with respect to controlledrelease of the active substance. The coating may be applied on singleunit dosage forms (e.g. tablets, capsules) or it may be applied on apolydepot dosage form or on its individual units.

Suitable coating materials are e.g. methylcellulose,hydroxypropylmethylcellulose, hydroxypropylcellulose, acrylic polymers,ethylcellulose, cellulose acetate phthalate, polyvinyl acetatephthalate, hydroxypropyl methylcellulose phthalate, polyvinylalcohol,sodium carboxymethylcellulose, cellulose acetate, cellulose acetatephthalate, gelatin, methacrylic acid copolymer, polyethylene glycol,shellac, sucrose, titanium dioxide, carnauba wax, microcrystalline wax,glyceryl monostearate, zein.

Plasticizers and other ingredients may be added in the coating material.The same or different active substance may also be added in the coatingmaterial.

In preferred embodiments of the invention, the solid dosage forms aredesigned to release tacrolimus and/or an analogue thereof in acontrolled manner. In the present context, the term “controlled manner”is intended to include all types of release which differ from therelease obtained from plain tablets. Thus, the term includes so-called“controlled release”, “modified release”, “sustained release”, “pulsedrelease”, “prolonged release”, burst release”, “slow release”, “extendedrelease”, as well as the terms “delayed release” and pH dependantrelease. However, a specific aspect of the invention relates to adelayed release composition or dosage form, which in this context isintended to denote a composition or dosage form that at the mostreleases 10% w/w of the active substance within the first 2 hours afteradministration and/or after start of a dissolution test employing adissolution medium having a pH of at the most about 3.

Modified Release Systems

A first modified release system includes matrix systems, in whichtacrolimus is embedded or dispersed in a matrix of another material thatserves to retard the release of tacrolimus into an aqueous environment(i.e., the luminal fluid of the GI tract). When tacrolimus is dispersedin a matrix of this sort, release of the drug takes place principallyfrom the surface of the matrix. Thus the drug is released from thesurface of a device, which incorporates the matrix after it diffusesthrough the matrix or when the surface of the device erodes, exposingthe drug. In some embodiments, both mechanisms can operatesimultaneously. The matrix systems may be large, i.e., tablet sized(about 1 cm), or small (<0.3 cm). The system may be unitary (e.g., abolus), may be divided by virtue of being composed of several sub-units(for example, several capsules which constitute a single dose) which areadministered substantially simultaneously, or may comprise a pluralityof particles, also denoted a multiparticulate. A multiparticulate canhave numerous formulation applications. For example, a multiparticulatemay be used as a powder for filling a capsule shell, or used per se formixing with food to ease the intake.

In a specific embodiment, a matrix multiparticulate, comprises aplurality of tacrolimus-containing particles, each particle comprisingtacrolimus and/or an analogue thereof e.g. in the form of a solidsolution/dispersion with one or more excipients selected to form amatrix capable of controlling the dissolution rate of the tacrolimusinto an aqueous medium. The matrix materials useful for this embodimentare generally hydrophobic materials such as waxes, some cellulosederivatives, or other hydrophobic polymers. If needed, the matrixmaterials may optionally be formulated with hydrophobic materials, whichcan be used as binders or as enhancers. Matrix materials useful for themanufacture of these dosage forms such as: ethylcellulose, waxes such asparaffin, modified vegetable oils, carnauba wax, hydrogenated castoroil, beeswax, and the like, as well as synthetic polymers such aspoly(vinyl chloride), poly(vinyl acetate), copolymers of vinyl acetateand ethylene, polystyrene, and the like. Water soluble or hydrophilicbinders or release modifying agents which can optionally be formulatedinto the matrix include hydrophilic polymers such as hydroxypropylcellulose (HPC), hydroxypropyl methyl cellulose (HPMC), methylcellulose, poly(N-vinyl-2-pyrrolidinone) (PVP), poly(ethylene oxide)(PEO), poly(vinyl alcohol) (PVA), xanthan gum, carrageenan, and othersuch natural and synthetic materials. In addition, materials, whichfunction as release-modifying agents include water-soluble materialssuch as sugars or salts. Preferred water-soluble materials includelactose, sucrose, glucose, and mannitol, as well as hydrophillicpolymers like e.g. HPC, HPMC, and PVP.

In a specific embodiment, a multiparticulate product is defined as beingprocessed by controlled agglomeration. In this case tacrolimus isdissolved or partly dissolved in a suitable meltable carrier and sprayedon carrier particles comprising the matrix substance. Suitable meltablecarriers are mentioned previously herein.

Alternatively, tacrolimus is dissolved in an organic solvent togetherwith the matrix substance and spray dried or applied to carrierparticles, cf. below. Solvents typically employed for the processinclude acetone, ethanol, isopropanol, ethyl acetate, and mixtures oftwo or more.

Once formed, tacrolimus matrix multiparticulates may be blended withcompressible excipients such as lactose, microcrystalline cellulose,dicalcium phosphate, and the like and the blend compressed to form atablet. Disintegrants such as sodium starch glycolate or crosslinkedpoly(vinyl pyrrolidone) are also usefully employed. Tablets prepared bythis method disintegrate when placed in an aqueous medium (such as theGI tract), thereby exposing the multiparticulate matrix, which releasestacrolimus therefrom.

In a further embodiment, the matrix system is in the form of ahydrophilic matrix tablet containing tacrolimus and/or an analoguethereof (e.g. in the form of a solid dispersion) as a multiparticulateproduct and an amount of hydrophilic polymer sufficient to provide auseful degree of control over the tacrolimus dissolution. Hydrophilicpolymers useful for forming the matrix include hydroxypropylmethylcellulose (HPMC), hydroxypropyl cellulose (HPC), poly(ethylene oxide),poly(vinyl alcohol), xanthan gum, carbomer, carrageenan, and zooglan. Apreferred material is HPMC. Other similar hydrophilic polymers may alsobe employed. In use, the hydrophilic material is swollen by, andeventually dissolves in, water. The tacrolimus is released both bydiffusion from the matrix and by erosion of the matrix. The tacrolimusdissolution rate of these hydrophilic matrix tablets may be controlledby the amount, molecular weight and gel strengths of the hydrophilicpolymer employed. In general, using a greater amount of the hydrophilicpolymer decreases the dissolution rate, as does using a higher molecularweight polymer. Using a lower molecular weight polymer normallyincreases the dissolution rate. A matrix tablet typically comprisesabout 20 to 90% by weight of tacrolimus and about 80 to 10% by weight ofpolymer.

A preferred matrix tablet comprises, by weight, about 30% to about 80%solid dispersion containing tacrolimus and/or an analogue thereof about15% to about 35% matrix former (such as, e.g., HPMC), 0% to about 35%lactose, 0% to about 20% microcrystalline cellulose, and about 0.25% toabout 2% lubricant (such as, e.g., magnesium stearate).

The matrix systems as a class often exhibit non-constant release of thedrug from the matrix. This result may be a consequence of the diffusivemechanism of drug release, and modifications to the geometry of thedosage form can be used with advantage to make the release rate of thedrug more constant.

A second class of tacrolimus controlled-release dosage forms of thisinvention includes membrane-moderated or reservoir systems. In thisclass, a reservoir of tacrolimus e.g. in a solid solution/dispersion asa multiparticulate product is surrounded by a rate-limiting membrane.The tacrolimus traverses the membrane by mass transport mechanisms wellknown in the art, including but not limited to dissolution in themembrane followed by diffusion across the membrane or diffusion throughliquid-filled pores within the membrane. These individual reservoirsystem dosage forms may be large, as in the case of a tablet containinga single large reservoir, or multiparticulate, as in the case of acapsule or poly-depot tablets containing a plurality of reservoirparticles, each individually coated with a membrane. The coating can benon-porous, yet permeable to tacrolimus (for example tacrolimus maydiffuse directly through the membrane), or it may be porous. As withother embodiments of this invention, the particular mechanism oftransport is not believed to be critical.

Sustained release coatings as known in the art may be employed tofabricate the membrane, especially polymer coatings, such as a celluloseester or ether, an acrylic polymer, or a mixture of polymers. Preferredmaterials include ethyl cellulose, cellulose acetate and celluloseacetate butyrate. The polymer may be applied as a solution in an organicsolvent or as an aqueous dispersion or latex. The coating operation maybe conducted in standard equipment such as a fluid bed coater, a Wurstercoater, or a rotary fluid bed coater.

If desired, the permeability of the coating may be adjusted by blendingof two or more materials. A particularly useful process for tailoringthe porosity of the coating comprises adding a pre-determined amount ofa finely-divided water-soluble material, such as sugars or salts orwater-soluble polymers to a solution or dispersion (e.g., an aqueouslatex) of the membrane-forming polymer to be used. When the dosage formis ingested into the aqueous medium of the GI tract, these water solublemembrane additives are leached out of the membrane, leaving pores whichfacilitate release of the drug. The membrane coating can also bemodified by the addition of plasticizers, as known in the art.

A particularly useful variation of the process for applying a membranecoating comprises dissolving the coating polymer in a mixture ofsolvents chosen such that as the coating dries, a phase inversion takesplace in the applied coating solution, resulting in a membrane with aporous structure.

In general, a support for mechanically strengthening the membrane is notrequired.

The morphology of the membrane is not of critical importance so long asthe permeability characteristics enumerated herein are met. The membranecan be amorphous or crystalline. It can have any category of morphologyproduced by any particular process and can be, for example, aninterracially-polymerized membrane (which comprises a thin rate-limitingskin on a porous support), a porous hydrophilic membrane, a poroushydrophobic membrane, a hydrogel membrane, an ionic membrane, and othersuch materials which are characterized by controlled permeability totacrolimus.

In one embodiment of the invention it is an aim to reduce the exposureof the upper GI tract to high concentrations of tacrolimus. Accordingly,suitable dosage forms includes those forms, which incorporate a specificdelay before the onset of controlled release of tacrolimus. An exemplaryembodiment can be illustrated by a tablet (or a particulate material)comprising a core containing tacrolimus coated with a first coating of apolymeric material of the type useful for sustained release oftacrolimus and a second coating of the type useful for delaying releaseof drugs when the dosage form is ingested. The first coating is appliedover and surrounds the tablet or individual particles. The secondcoating is applied over and surrounds the first coating.

A tablet can be prepared by techniques well known in the art andcontains a therapeutically useful amount of tacrolimus plus suchexcipients as are necessary to form the tablet by such techniques.

The first coating may be a sustained release coating as known in theart, especially polymer coatings, to fabricate the membrane, aspreviously discussed for reservoir systems, or it could be a controlledrelease matrix core, which are coated a second time with a delayedrelease material.

Materials useful for preparing the second coating on the tablet includepolymers known in the art as enteric coatings for delayed-release ofpharmaceuticals. These most commonly are pH-sensitive materials such ascellulose acetate phthalate, cellulose acetate trimellitate,hydroxypropyl methyl cellulose phthalate, poly(vinyl acetate phthalate),and acrylic copolymers such as Eudragit L-100 (Röhm Pharma) and relatedmaterials, as more fully detailed below under “Delayed Release”. Thethickness of the delayed-release coating is adjusted to give the desireddelay property. In general, thicker coatings are more resistant toerosion and, consequently, yield a longer and more effective delay.Preferred coatings range from about 30 μm in thickness to about 3 mm inthickness.

When using a hydrophobic matrix material like glyceryl monostearate, nodelay coating is necessary. The tablet will not release tacrolimus untilan area of enzymatic degradation has been reached, more specificallyafter the duodenum.

When ingested, the twice-coated tablet passes through the stomach, wherethe second coating prevents release of the tacrolimus under the acidicconditions prevalent there. When the tablet passes out of the stomachand into the small intestine, where the pH is higher, the second coatingerodes or dissolves according to the physicochemical properties of thechosen material. Upon erosion or dissolution of the second coating, thefirst coating prevents immediate or rapid release of the tacrolimus andmodulates the release so as to prevent the production of high peakconcentrations, thereby minimizing side-effects.

A further preferred embodiment comprises a multiparticulate wherein eachparticle is dual coated as described above for tablets, first with apolymer designed to yield sustained release of the tacrolimus and thencoated with a polymer designed to delay onset of release in theenvironment of the GI tract when the dosage form is ingested.

The rate of tacrolimus release from the sustained-release-coatedmultiparticulates (i.e., the multiparticulates before they receive thedelayed-release coating) and methods of modifying the coating are alsocontrolled by the factors previously discussed for reservoir systemtacrolimus multiparticulates.

The second membrane or coating for dual coated multiparticulates is adelayed-release coating which is applied over the firstsustained-release coating, as disclosed above for tablets, and may beformed from the same materials. It should be noted that the use of theso-called “enteric” materials to practice this embodiment differssignificantly from their use to produce conventional enteric dosageforms. With conventional enteric forms, the object is to delay releaseof the drug until the dosage form has passed the stomach and then todeliver the dose in the duodenum. Dosing of tacrolimus directly andcompletely to the duodenum may be undesirable, however, due to the sideeffects sought to be minimized or avoided by this invention. Therefore,if conventional enteric polymers are to be used to practice thisembodiment, it may be necessary to apply them significantly more thicklythan in conventional practice, in order to delay drug release until thedosage form reaches the lower GI tract. However, it is also possible toeffect a sustained or controlled delivery of tacrolimus after thedelayed-release coating has dissolved or eroded, therefore the benefitsof this embodiment may be realized with a proper combination ofdelayed-release character with sustained-release character, and thedelayed-release part alone may or may not necessarily conform to USPenteric criteria. The thickness of the delayed-release coating isadjusted to give the desired delay property. In general, thickercoatings are more resistant to erosion and, consequently, yield a longerdelay.

A first delayed release embodiment according to the invention is a“pH-dependent coated dosage form” such as, e.g., a tablet or a capsule.In the case of a tablet it comprises a tablet core comprising tacrolimuse.g. in a solid solution/dispersion as a multiparticulate product, acontrolled release matrix of e.g. HPMC, a disintegrant, a lubricant, andone or more pharmaceutical carriers, such core being coated with amaterial, preferably a polymer, which is substantially insoluble andimpermeable at the pH of the stomach, and which is more soluble andpermeable at the pH of the small intestine. Preferably, the coatingpolymer is substantially insoluble and impermeable at pH<5.0, andwater-soluble at pH>5.0. The tablet core may be coated with an amount ofpolymer sufficient to assure that substantially no release of tacrolimusfrom the dosage form occurs until the dosage form has exited the stomachand has resided in the small intestine for about 15 minutes or greater,preferably about 30 minutes or greater, thus assuring that minimaltacrolimus is released in the duodenum. Mixtures of a pH-sensitivepolymer with a water-insoluble polymer may also be employed. Tablets arecoated with an amount of polymer comprising from about 10% to about 80%of the weight of the tacrolimus-containing tablet core. Preferredtablets are coated with an amount of polymer comprising about 15% toabout 50% of the weight of the tacrolimus tablet core.

pH-sensitive polymers which are very insoluble and impermeable at the pHof the stomach, but which are more soluble and permeable at the pH ofthe small intestine and colon include polyacrylamides, phthalatederivatives such as acid phthalates of carbohydrates, amylose acetatephthalate, cellulose acetate phthalate, other cellulose esterphthalates, cellulose ether phthalates, hydroxypropylcellulosephthalate, hydroxypropylethylcellulose phthalate,hydroxypropylmethylcellulose phthalate, methylcellulose phthalate,polyvinyl acetate phthalate, polyvinyl acetate hydrogen phthalate,sodium cellulose acetate phthalate, starch acid phthalate,styrene-maleic acid dibutyl phthalate copolymer, styrene-maleic acidpolyvinylacetate phthalate copolymer, styrene and maleic acidcopolymers, polyacrylic acid derivatives such as acrylic acid andacrylic ester copolymers, polymethacrylic acid and esters thereof, polyacrylic methacrylic acid copolymers, shellac, and vinyl acetate andcrotonic acid copolymers.

Preferred pH-sensitive polymers include shellac; phthalate derivatives,particularly cellulose acetate phthalate, polyvinylacetate phthalate,and hydroxypropylmethylcellulose phthalate; polyacrylic acidderivatives, particularly polymethyl methacrylate blended with acrylicacid and acrylic ester copolymers; and vinyl acetate and crotonic acidcopolymers.

The delay time before release of tacrolimus, after the “pH-dependentcoated tablet” dosage form has exited the stomach, may be controlled bychoice of the relative amounts of Eudragit-L® and Eudragit-S® in thecoating, and by choice of the coating thickness. Eudragit-L® filmsdissolve above pH 6.0, and Eudragit-Se films dissolve above 7.0, andmixtures dissolve at intermediate pH's. Since the pH of the duodenum isapproximately 6.0 and the pH of the colon is approximately 7.0, coatingscomposed of mixtures of Eudragit-Le and Eudragit-S® provide protectionof the duodenum from tacrolimus. If it is desired to delay release oftacrolimus until the tacrolimus-containing “pH-dependent coated tablet”has reached the colon, Eudragit-S® may be used as the coating material,as described by Dew et al. (Br. J. Clin. Pharmac. 14 (1982) 405-408). Inorder to delay the release of tacrolimus for about 15 minutes or more,preferably 30 minutes or more, after the dosage form has exited thestomach, preferred coatings comprise from about 9:1 to about 1:9Eudragit-L®/Eudragit-S®, more preferably from about 9:1 to about 1:4Eudragit-L®/Eudragit-S®. The coating may comprise from about 3% to about70% of the weight of the uncoated tablet core. Preferably, the coatingcomprises from about 5% to about 50% of the weight of the tablet core.

Uses

The solid dispersion and/or solution of the invention or thepharmaceutical composition of the invention may be used in thepreparation of an solid oral dosage form such as tablets, capsules orsachets; or for the preparation of granules, pellets microspheres ornanoparticles.

Preferably, the solid dispersion or solid solution is used in thepreparation of an immediate release solid dosage form or a delayedrelease solid dosage form.

Other uses of the solid dispersion or solid solution of the invention isfor the preparation of a topical dosage form.

A further advantage of a composition of the present invention is thepossibility of obtaining an effective therapeutic response with adecreased dosage compared to traditional oral treatment. Thus it iscontemplated that the solid dosage form of the invention, when orallyadministered to a mammal in need thereof in a dose that is at the mostabout 85% w/w such as, e.g., at the most about 80% w/w, at the mostabout 75%, at the most about 70% w/w, at the most about 65% w/w, at themost about 60% w/w, at the most about 55% why or at the most about 50%w/w of the dose of tacrolimus administered in the form of Prograf® or asimilar commercially available tacrolimus-containing product, isessentially bioequivalent with Prograf® or a similar commerciallyavailable tacrolimus-containing product.

Any of the tacrolimus-containing dosage forms, compositions, dispersionsor solutions of the invention may improved treatment of conditions thatrespond to tacrolimus treatment.

Tacrolimus is indicated (or has been suggested) for the treatment ofdiseases such as, e.g., rejection reactions by transplantation of organsor tissues such as the heart, kidney, liver, bone marrow, skin, cornea,lung, pancreas, small intestine, limb, muscle, nerve, intervertebraldisc, trachea, myoblast, cartilage, etc.; graft-versus-host reactionsfollowing bone marrow transplantation; autoimmune diseases such asrheumatoid arthritis, systemic lupus erythematosus, Hashimoto'sthyroiditis, multiple sclerosis, myasthenia gravis, type I diabetes,etc.; infections caused by pathogenic microorganisms (e.g. Aspergillusfumigatus, Fusarium oxysporum, Trichophyton asteroides, etc.);inflammatory or hyperproliferative skin diseases or cutaneousmanifestations of immunologically mediated diseases (e.g. psoriasis,atopic dermatitis, contact dermatitis, eczematoid dermatitis, seborrheicdermatitis, lichen planus, pemphigus, bullous pemphigoid, epidermolysisbullosa, urticaria, angioedema, vasculitides, erythema, dermaleosinophilia, lupus erythematosus, acne, and alopecia areata);autoimmune diseases of the eye (e.g. keratoconjunctivitis, vernalconjunctivitis, uveitis associated with Behcet's disease, keratitis,herpetic keratitis, conical keratitis, corneal epithelial dystrophy,keratoleukoma, ocular premphigus, Mooren's ulcer, scleritis, Graves'ophthalmopathy, Vogt-Koyanagi-Harada syndrome, keratoconjunctivitissicca (dry eye), phlyctenule, iridocyclitis, sarcoidosis, endocrineophthalmopathy, etc.); reversible obstructive airways diseases [asthma(e.g. bronchial asthma, allergic asthma, intrinsic asthma, extrinsicasthma, and dust asthma), particularly chronic or inveterate asthma(e.g. late asthma and airway hyper-responsiveness) bronchitis, etc.;mucosal or vascular inflammations (e.g. gastric ulcer, ischemic orthrombotic vascular injury, ischemic bowel diseases, enteritis,necrotizing enterocolitis, intestinal damages associated with thermalburns, leukotriene B4-mediated diseases); intestinalinflammations/allergies (e.g. coeliac diseases, proctitis, eosinophilicgastroenteritis, mastocytosis, Crohn's disease and ulcerative colitis);food-related allergic diseases with symptomatic manifestation remotefrom the gastrointestinal tract (e.g. migraine, rhinitis and eczema);renal diseases (e.g. intestinal nephritis, Goodpasture's syndrome,hemolytic uremic syndrome, and diabetic nephropathy); nervous diseases(e.g. multiple myositis, Guillain-Barre syndrome, Meniere's disease,multiple neuritis, solitary neuritis, cerebral infarction, Alzheimer'sdiseases Parkinson's diseases, amyotrophic lateral sclerosis (ALS) andradiculopathy); cerebral ischemic disease (e.g., head injury, hemorrhagein brain (e.g., subarachnoid hemorrhage, intracerebral hemorrhage),cerebral thrombosis, cerebral embolism, cardiac arrest, stroke,transient ischemic attack (TIA), hypertensive encephalopathy, cerebralinfarction); endocrine diseases (e.g. hyperthyroidism, and Basedow'sdisease); hematic diseases (e.g. pure red cell aplasia, aplastic anemia,hypoplastic anemia, idiopathic thrombocytopenic purpura, autoimmunehemolytic anemia, agranulocytosis, pernicious anemia, megaloblasticanemia, and anerythroplasia); bone diseases (e.g. osteoporosis);respiratory diseases (e.g. sarcoidosis, pulmonary fibrosis, andidiopathic interstitial pneumonia); skin diseases (e.g. dermatomyositis,leukoderma vulgaris, ichthyosis vulgaris, photosensitivity, andcutaneous T-cell lymphoma); circulatory diseases (e.g. arteriosclerosis,atherosclerosis, aortitis syndrome, polyarteritis nodosa, andmyocardosis); collagen diseases (e.g. scleroderma, Wegener's granuloma,and Sjogren's syndrome); adiposis; eosinophilic fascitis; periodontaldiseases (e.g. damage to gingiva, periodontium, alveolar bone orsubstantia ossea dentis); nephrotic syndrome (e.g. glomerulonephritis);male pattern alopecia, alopecia senile; muscular dystrophy; pyoderma andSezary syndrome; chromosome abnormality-associated diseases (e.g. Down'ssyndrome); Addison's disease; active oxygen-mediated diseases [e.g.organ injury (e.g. ischemic circulation disorders of organs (e.g. heart,liver, kidney, digestive tract, etc.) associated with preservation,transplantation, or ischemic diseases (e.g. thrombosis, cardialinfarction, etc.)); intestinal diseases (e.g. endotoxin shock,pseudomembranous colitis, and drug- or radiation-induced colitis); renaldiseases (e.g. ischemic acute renal insufficiency, chronic renalfailure); pulmonary diseases (e.g. toxicosis caused by pulmonary oxygenor drugs (e.g. paracort, bleomycin, etc.), lung cancer, and pulmonaryemphysema); ocular diseases (e.g. cataracta, iron-storage disease(siderosis bulbi), retinitis, pigmentosa, senile plaques, vitreousscarring, corneal alkali burn); dermatitis (e.g. erythema multiforme,linear immunoglobulin A bullous dermatitis, cement dermatitis); andother diseases (e.g. gingivitis, periodontitis, sepsis, pancreatitis,and diseases caused by environmental pollution (e.g. air pollution),aging, carcinogen, metastasis of carcinoma, and hypobaropathy)];diseases caused by histamine release or leukotriene C4 release;restenosis of coronary artery following angioplasty and prevention ofpostsurgical adhesions; autoimmune diseases and inflammatory conditions(e.g., primary mucosal edema, autoimmune atrophic gastritis, prematuremenopause, male sterility, juvenile diabetes mellitus, pemphigusvulgaris, pemphigoid, sympathetic ophthalmitis, lens-induced uveitis,idiopathic leukopenia, active chronic hepatitis, idiopathic cirrhosis,discoid lupus erythematosus, autoimmune orchitis, arthritis (e.g.arthritis deformans), or polychondritis); Human Immunodeficiency Virus(HIV) infection, AIDS; allergic conjunctivitis; hypertrophic cicatrixand keloid due to trauma, burn, or surgery.

In addition, tricyclic macrolides like e.g. tacrolimus have liverregenerating activity and/or activities of stimulating hypertrophy andhyperplasia of hepatocytes. Therefore, the pharmaceutical composition ofthe present invention is useful for increasing the effect of the therapyand/or prophylaxis of liver diseases [e.g. immunogenic diseases (e.g.chronic autoimmune liver diseases such as autoimmune hepatic diseases,primary biliary cirrhosis or sclerosing cholangitis), partial liverresection, acute liver necrosis (e.g. necrosis caused by toxins, viralhepatitis, shock, or anoxia), hepatitis B, non-A non-B hepatitis,hepatocirrhosis, and hepatic failure (e.g. fulminant hepatitis,late-onset hepatitis and “acute-on-chronic” liver failure (acute liverfailure on chronic liver diseases))].

Furthermore, a composition of the present invention is useful forincreasing the effect of the prevention and/or treatment of variousdiseases because of the useful pharmacological activity of the tricyclicmacrolides, such as augmenting activity of chemotherapeutic effect,activity of cytomegalovirus infection, anti-inflammatory activity,inhibiting activity against peptidyl-prolyl isomerase or rotamase,antimalarial activity, antitumor activity and so on.

Materials and Methods

Materials

Tacrolimus (supplied by Eurotrade); batch no RD 03-111

Lactose monohydrate 200 mesh (from DMV)

Granulated silicium oxide, Aeroperl® 300, (Degussa)

Polyethylene glycol 6000, Pluracol® E6000 (from BASF)

Poloxamer 188, Pluronic® F-68 (from BASF)

Glyceryl monostearate, Rylo® MD50, (from Danisco Cultor), Ph. Eur.;batch no. 4010056276

Avicel PH200 (microcrystalline cellulose) (from FMC)

Lactose DCL 11 (from DMV)

Magnesium stearate

Croscarmellose sodium, Ac-Di-Sol® (from FMC)

Eudragit® L30D.55 (from Degussa)

Triethyl citrate (from Merck)

Anti-foam emulsion (from Unikem)

Micro talc

HPMC refers to Metolose 90SH (type 2910, 2208) or Metolose 60SH (type2910) from ShinEtsu available in various degrees of polymerization(viscosity 3-100,000 cP).

Tablets, capsules or granules might be enteric coated with differenttypes of polymers such as hydroxypropylmethylcellulose acetate succinate(Aqoat), cellulose acetate phthalate CAP, hydroxypropylmethylcellulosephtalate HPMCP or methacrylic acid copolymers such as Eudragit L30D,Eudragit 100/S, Eudragit 100/L.

Comparison Prior Art Tacrolimus Formulation for In Vivo Studies:

Prograt® Hard Gelatin Capsules, manufactured by Fujisawa Ireland Ltd.

Ingredients mg Tacrolimus, anhydr. 1.0 Gelatin 6.9 Hypromellose 1.0Lactose monohydrate 24.7 Magnesium stearate 0.3 Shellac q.s. Soybeanlecitine q.s. Iron oxide red (E172) q.s. Titanium dioxide (E171) q.s.Dimeticone (E900) q.s.MethodsDetermination of Weight Variation

The tablets prepared in the Examples herein were subjected to a test forweight variation performed in accordance with Ph. Eur.

Determination of Average Tablet Hardness

The tablets prepared in the Examples herein were subjected to at testfor tablet hardness employing Schleuniger Model 6D apparatus andperformed in accordance with the general instructions for the apparatus.

Determination of Disintegration Time

The time for a tablet to disintegrate, i.e. to decompose into particlesor agglomerates, was determined in accordance with Ph. Eur.

Determination of Geometric Weight Mean Diameter d_(gw)

The geometric weight mean diameter was determined by employment of amethod of laser diffraction dispersing the particulate material obtained(or the starting material) in air. The measurements were performed at 1bar dispersive pressure in Sympatec Helos equipment, which records thedistribution of the equivalent spherical diameter. This distribution isfitted to a log normal volume-size distribution.

When used herein, “geometric weight mean diameter” means the meandiameter of the log normal volume-size distribution.

In Vitro Dissolution Tests

The following test methods were applies to the compositions and dosageforms of the present invention.

Test 1:

In vitro dissolution test according to USP Method A, delayed releasearticles (USP paddle method; rotation speed: 50 rpm; 37° C.; after 2hours in acidic medium, the medium is changed to phosphate buffer pH6.8.).

Test 2:

In vitro dissolution test in aqueous dissolution medium adjusted to pH4.5 (900 ml water with 0.005% HPC (hydroxypropylcellulose) adjusted topH4.5; 37° C.; USP Paddle method; rotation speed: 50 rpm).

In Vivo Studies in Beagle Dogs

In vivo studies with the purpose of determining the bioavailability ofthe compositions of the present invention relative to thebioavailability of the commercially available tacrolimus product, i.e.Prograf®, was performed using Beagle dogs.

The experimental work was performed in Denmark using male Beagle dogseach having a body weight of 12-18 kg (starting weight). The studieswere conducted as open, non-randomised, cross-over studies. Each animalwas its own control. The dogs were premedicated with Primperan inj. 5mg/ml (anti-emetica) and an oral dose of 0.5 to 4 mg of tacrolimus wasadministered.

The dogs were fasted for 10 hours prior to dosing (water ad libitum) andwere fed 5 hours after dosing (water ad libitum). Each dog was dosedwith the specified dose of tacrolimus without taking the weight of thedog into consideration.

Blood samples were collected at vena jugularis externa at the followingpoints of time:

Pre-dose, 1, 1.5, 2, 3, 4, 6, 8, 12 and 24 hours after dosing. 4 ml ofblood were collected, mixed with EDTA, and the samples were frozen (−80°C.). The blood samples were analyzed using on-line extraction LC/MS andresults were given in ng/mL.

The determined full blood concentration profiles of tacrolimus weretreated using the Pharmacokinetic softwear WinNonlin®, (Pharsight,Calif.; USA) to calculate the pharmacokinetic parameters. All data aredose adjusted.

In Vivo Studies in Göttingen Mini-Pigs

In vivo studies with the purpose of determining the bioavailability ofthe compositions of the present invention relative to thebioavailability of the commercially available tacrolimus product, i.e.Prograf®, was performed using Göttingen mini-pigs.

The experimental work was performed in Denmark using female mini-pigshaving a body weight of 15-18 kg (starting weight). The studies wereconducted as open, non-randomised, cross-over studies. Each animal wasits own control. An oral dose of 1 mg of tacrolimus was administered.

The mini-pigs were fasted for 24 hours prior to dosing (water adlibitum), the mini-pigs were allowed to eat 24 hours after dosing. Eachmini-pig was dosed with the specified dose of Tacrolimus without takingthe weight of the mini-pig into consideration.

Blood samples were collected at vena jugularis extema at the followingpoints of time:

Pre-dose, 0.5, 1, 1.5, 2, 3, 4, 6, 8, 12 and 24 hours after dosing. 4 mlof blood were collected, mixed with EDTA, and the samples were frozen(−80° C.). The blood samples were analyzed using on-line extractionLC/MS and results were given in ng/mL.

The determined full blood concentration profiles of tacrolimus weretreated using the Pharmacokinetic softwear WinNonlin®, (Pharsight,Calif.; USA) to calculate the pharmacokinetic parameters. All data aredose adjusted.

The following examples serve the purpose of illustration of theinvention and are not intended to limiting the scope of the presentinvention.

Pharmaceutical compositions and dosage forms of the invention areexemplified in examples 1-16. Results of in vitro dissolution tests ofcompositions and dosage forms of the invention are found in example 17.Results of in vivo comparison studies in Beagle dogs (blood plasmaconcentration) are found in example 18, and results of in vivocomparison studies in Göttingen mini-pigs (blood plasma concentration)are found in example 19.

Example 1 Modified Release Polydepot Capsule Based on SwellingHydrocolloid Matrix of Hydroxypropylcellulose

Substance % mg Tacrolimus 0.50 1.00 HPMC 20.00 40.00 Lactose 200 mesh30.00 60.00 PEG 6000 34.65 69.30 Poloxamer 188 14.85 29.70 Total 100.00200.00

Tacrolimus was dissolved in polyethylene glycol 6000 and poloxamer 188(70:30 w/w ratio) at 70° C. The solution was sprayed on a mixture of 150g lactose and 100 g HPMC in a fluid bed Strea-1. The granular productwas sieved through sieve 0.7 mm and filled into hard gelatine capsules(200 mg).

Example 2 Modified Release Polydepot Capsule Based on SwellingHydrocolloid Matrix of Hydroxypropylcellulose

Substance % mg Tacrolimus 0.50 1.00 HPMC 2910 3 cp 20.00 40.00 Lactose200 mesh 30.00 60.00 Glyceryl monostearate 49.50 99.00 Total 100.00200.00

Tacrolimus was dissolved in glyceryl monostearate at 70° C. The solutionwas sprayed on a mixture of 150 g lactose and 100 g HPMC in a fluid bedStrea-1. The granular product was sieved through sieve 0.7 mm and filledinto hard gelatine capsules (200 mg).

Example 3 Modified Release Matrix Tablet Based on Swelling HydrocolloidMatrix of Hydroxypropylcellulose

Substance % mg Tacrolimus 0.50 1.00 HPMC 19.90 40.00 Lactose 200 mesh29.85 60.00 PEG 6000 34.48 69.30 Poloxamer 188 14.78 29.70 Magnesiumstearate 0.50 1.01 Total 100.00 201.01

Tacrolimus was dissolved in polyethylene glycol 6000 and poloxamer 188(70:30 w/w ratio) at 70° C. The solution was sprayed onto 250 g lactosein a fluid bed Strea-1. The resulting granular product was sievedthrough sieve 0.7 mm and blended with HPMC and magnesium stearate for0.5 min in a Turbula mixer.

The mixture was compressed into 8 mm tablets of 1 mg active ingredient(200 mg tablet) with compound cup shaped.

Mean disintegration time: 20 min. Hardness: 45 N

Example 4 Modified Release Matrix Tablet Based on Lipophilic Matrix ofGlyceryl Monostearate

Substance % mg Tacrolimus 0.50 1.00 Lactose 200 mesh 49.75 100.00Glyceryl monostearate 49.25 99.00 Magnesium stearate 0.50 1.01 100.00201.01

Tacrolimus was dissolved in glyceryl monostearate at 70° C. The solutionwas sprayed onto 250 g lactose in a fluid bed Strea-1. The granularproduct was sieved through sieve 0.7 mm and blended with magnesiumstearate for 0.5 minutes in a Turbula mixer.

The resulting mixture was compressed into 8 mm tablets of 1 mg activeingredient (200 mg tablet) with compound cup shape.

Mean disintegration time: 20 min. Hardness: 45 N

Example 5 Modified Release Polydepot Capsule Based on Lipophilic Matrixof Glyceryl Monostearate

Substance % mg Tacrolimus 0.50 1.00 Lactose 200 mesh 49.75 100.00Glyceryl monostearate 49.25 99.00 Magnesium stearate 0.50 1.01 100.00201.01

Tacrolimus was dissolved in glyceryl monostearate at 70° C. The solutionwas sprayed onto 250 g lactose in a fluid bed Strea-1. The granularproduct was sieved through sieve 0.7 mm and filled into hard gelatinecapsules (200 mg).

Example 6 Modified Release Polydepot Tablet Based on Lipophilic Matrixof Gelucire® 44/14

Substance % mg Tacrolimus 0.50 1.00 Aeroperl ® 300 49.75 100.00Gelucire ® 44/14 49.25 99.00 Magnesium stearate 0.50 1.01 100.00 201.01

Tacrolimus was dissolved in Gelucire® at 70° C. The solution was sprayedonto 250 g Aeroperl® in a fluid bed Strea-1. The granular product wassieved through sieve 0.7 mm and filled into hard gelatine capsules (200mg).

The resulting granulate was compressed into 8 mm tablets of 1 mg activeingredient (tablet weight 200 mg). Tablets were cup shaped.

Mean disintegration time: 25 minutes. Hardness: 43 N.

Example 7 Enteric Coating

Capsules and tablets of examples 1, 2, 3, 5 and 6 were subsequentlycoated with the following enteric coating in order to obtain a delayedrelease of active ingredient after administration.

Ingredients % Eudragit ® L30D 40 Purified water 52 Triethylacetylcitrate 1.8 Anti-foam emulsion 0.2 Talc 6 Total 100

The coating suspension was prepared by mixing triethyl acetylcitrate,antifoam emulsion and purified water in Ultra Turrax apparatus at 9500rpm for 30 min. After 1 minute talc was added. The mixture was passedthrough sieve no. 300 and stirred by a magnet stirrer. Eudragit waspassed through sieve no. 300 and added the mixture, which was stirredfor 5 minutes.

The process conditions of the coating process were the following aninlet temperature of 40° C., an outlet temperature of 31° C., air inletof 140 cbm per hour and a coating time of approx. 50 minutes (300 g ofcoating material). Approx. 400 g of tablets, or 200 g of capsules werecoated.

The film coated tablets and capsules were cured for 48 hours at 30° C.before dissolution testing.

Example 8 Modified Release Matrix Tablet Based on Lipofilic Matrix ofGlycerol Monostearate

Substance % mg Tacrolimus 0.95 2.00 HPMC, Pharmacoat 606 6.75 14.29Lactose monohydrate, lactose 125 mesh 6.75 14.29 Glycerol monostearate ®Rylo ® MD50 30.56 64.67 Magnesium Stearate 0.5 1.06 Talc 4.5 9.52Lactose monohydrate, Pharmatose DCL 14 50.00 105.8 100.00 211.64

Tacrolimus was dissolved in glycerol monostearate at a temperature above80° C. The solution was sprayed by feed unit Phast FS1.7 onto 60 glactose and 60 g HPMC in a fluid bed Phast FB100. The granular productwas hardened in a heating oven for 4 hours at 50° C. The resultinggranular product was sifted through sieve 0.71 mm and blended withlactose for 3 minutes in a Turbula mixer.

Magnesium stearate and talc was sifted through sieve no. 300 and mixedin a Turbula mixer for 3 minutes. The granulate was mixed with themixture of magnesium stearate and talc (1:9) for 0.5 minutes in aTurbula mixer.

The final mixture was compressed into 8 mm tablets of 2 mg activeingredient (210 mg tablet) with compound cup shape.

Mean disintegration time: 2 hours. Hardness: 50 N

Example 9 Enteric Coated Tablet with Core Based on PEG 6000/Poloxamer188 and Enteric Coating Based on Eudragit L30D 55

Tablet Core Composition:

Substance % mg Tacrolimus 1.98 2.00 Lactose monohydrate, Lactose 200mesh 40.50 40.91 PEG 6000 33.26 33.60 Poloxamer 188, Lutrol 68 14.4014.40 Magnesium Stearate 0.50 0.51 Talc 4.50 4.55 Croscarmellose sodium,Ac-di-sol 5.00 5.05 100.00 101.01

The tacrolimus tablet core was produced by dissolving in PEG 6000 at atemperature above 80° C. Poloxamer 188 was added, and the solution washeated to a temperature above 80° C. The solution was sprayed by feedunit Phast FS1.7 on 200 g lactose monohydrate in a fluid bed PhastFB100. The resulting granulate was sifted through a Comill sieve 1397,4500 rpm, and blended with croscarmellose sodium for 3 minutes in aTurbula mixer.

Magnesium stearate and talc was sifted through sieve no. 300 and mixedin a Turbula mixer for 3 minutes. The granulate was mixed with magnesiumstearate and talc (1:9) for 0.5 minutes in a Turbula mixer.

The resulting mixture was compressed into 6 mm tablets of 2 mg activeingredient (100 mg tablet) with compound cup shape.

Mean disintegration time: 7 minutes. Hardness: 65 N

Enteric Coating:

The enteric coating is based on an acrylic polymer Eudragit L30D-55.Eudragit L30D is supplied as an aqueous latex suspension creating awater insoluble film when the water is evaporated during coating. Thepolymer is insoluble at pH-values below 5.0 and readily soluble atpH-values over 6.0. The film coating composition is:

Substance w/w % Eudragit L30D-55 40 Water 52 Triethyl citrate 1.8Anti-foam emulsion 0.2 Talc (micro) 6 Total 100

The amount of applied film polymer (Eudragit) was based on a calculationof mg film polymer per cm² tablet surface. The thickness of the entericcoating was 80 μm. A verification of the film thickness applied wasbased on measuring the increase in tablet height with a digitalmicrometer. The film coating process was performed in a Phast FB100fluid bed equipped with a Wurster like insert. The process conditionswere: Inlet air temperature 50° C.; Inlet air flow 100 m³ per hour,Product temperature 38° C.; Feed rate 15 g/min.

After coating proper film formation requires curing of the coatedtablets ie. 30° C. in 48 hours in an oven. Alternatively the coatedtablets more efficiently could be cured at 40° C. in 24 hours.

Example 10 Controlled Release PEG 6000/Poloxamer 188 Tablet Based on aHPMC Matrix

Tablet Composition:

Substance % mg Tacrolimus 1.21 2.00 Lactose monohydrate, Lactose 200mesh 24.75 40.91 PEG 6000 20.33 33.60 Poloxamer 188, Lutrol 68 8.7114.40 Magnesium Stearate 0.50 0.83 Talc 4.50 7.44 Hydroxypropylmethylcellulose, Metolose 90SH 15000 40.00 66.12 100.00 165.29

Tacrolimus was dissolved in PEG 6000 at a temperature above 80° C.Poloxamer 188 is added and the solution is heated to a temperature above80° C. The solution is sprayed by feed unit Phast FS1.7 on 200 g lactosemonohydrate in a fluid bed Phast FB100. The granular product is sievedthrough a Comill, sieve 1397, 4500 rpm, and blended with Hydroxypropylmethylcellulose for 3 min in a Turbula mixer.

Magnesium stearate and talc is sifted through sieve 300 and mixed in aTurbola mixer for 3 min. The granulate is mixed with MagnesiumStearate:Talc (1:9) for 0.5 min in a Turbula mixer.

The mixture is compressed into 8 mm tablets with strength of 2 mg (165mg tablet with compound cup shape).

Mean disintegration time: 2 hours 34 minutes, Hardness: 50 N

Example 11 Enteric Coated Tablet Formulation Wet Granulation andEnteric-Coated Tablets

Tablet Composition:

Ingredient mg Tacrolimus 2 Lactose 80 Sodium lauryl sulfate 10 KollidonVA64 3 Avicel PH200 30 Magnesium stearate 0.5 Total 125.5

The tablet formulation was based on wet granulation in a high shearmixer Pellmix ⅛. 16 g Micronized tacrolimus was mixed with 640 g lactose125 mesh and 80 g natrium lauryl sulfate in the high shear mixer. A 15%aqueous solution of binder Kolllidon VA64 was pumped to the mixture atan impeller speed of 500 rpm at a feed rate of 20 g/min. andsubsequently kneaded for 3 minutes at the equal speed. The granulate wasdried in a tray dryer and sieved through sieve size 0.7 mm.

The granulate was mixed with 240 g Avicel PH200 for 3 minutes and forand after addition of 4 g magnesium stearate for further 0.5 minute. Themixture was compressed into tablets on a single punch tabletting machineDiaf TM20.

Tablet diameter: 6 mm. Tablet shape: round, compound cup.

The tablets were subsequently coated with an enteric coating of acrylictype as described in example 9.

The amount of applied film polymer (Eudragit) should be based on acalculation of mg film polymer per cm² tablet surface. The thickness ofthe enteric coating should be 50-80 μm. A verification of the filmthickness applied is based on measuring the increase in tablet heightwith a digital micrometer. The film coating process is performed in aStre-1 fluid bed equipped with a Wurster insert at the following processconditions:

Process parameter Process value Product load, g 400 Inlet airtemperature, ° C. 40 Inlet air flow, m³ per hour 140 Outlet airtemperature, ° C. 31 Feed rate g/min 5

After coating, proper film formation requires curing of the coatedtablets, i.e. 30° C. in 48 hours in an oven. Alternatively the coatedtablets more efficiently could be cured at 40° C. for 24 hours.

Example 12 Controlled Release Tablet Formulation Based on Eroding HPMCMatrix

HPMC added as part of the extragranular phase. Wet granulation.

Tablet Composition:

Ingredients mg Tacrolimus 2 Lactose 80 Sodium lauryl sulfate 10 KollidonVA64 3 Avicel PH200 30 Metolose SH 90 60 Magnesium stearate 1 Total 186

The tablet formulation was based on wet granulation in a high shearmixer Pellmix ⅛. 16 g Micronized tacrolimus was mixed with 640 g lactose125 mesh and 80 g natrium lauryl sulfate in the high shear mixer. A 15%aqueous solution of binder Kolllidon VA64 was pumped to the mixture atan impeller speed of 500 rpm at a feed rate of 20 g/min and subsequentlykneaded for 3 minutes at equal impeller speed. The granulate was driedin a tray dryer and sieved through sieve size 0.7 mm.

The granulate was mixed with 240 g Avicel PH200 and 480 ghydroxypropylmethylcellulose Metolose SH 90 100 cP for 3 minutes and forand after addition of 8 g magnesium stearate for further 0.5 minute. Themixture was compressed into tablets on a single punch tabletting machineDiaf TM20.

Tablet diameter 7 mm. Tablet shape: round, compound cup.

Example 13 Controlled Release Tablet Formulation Based on Eroding HPMCMatrix

HPMC added as part of the intragranular phase. Wet granulation.

Tablet Composition:

Ingredient mg Tacrolimus 2 Lactose 80 Sodium lauryl sulfate 10 MetoloseSH 90 80 Avicel PH200 60 Magnesium stearate 2 Total 234

The tablet formulation was based on wet granulation in a high shearmixer Pellmix ⅛. 16 g Micronized tacrolimus was mixed with 640 g lactose125 mesh and 80 g natrium lauryl sulfate and 640 ghydroxypropylmethylcellulose Metolose SH 90 15.000 cP? in the high shearmixer. Purified water was pumped to the mixture at an impeller speed of500 rpm at a feed rate of 20 g/min. and subsequently kneaded for 3minutes. The granulate was dried in a tray dryer and sieved throughsieve size 0.7 mm.

The granulate was mixed with 480 g Avicel PH200 for 3 minutes and forand after addition of 16 g magnesium stearate for further 0.5 minute.The mixture was compressed into tablets on a single punch tablettingmachine Diaf TM20.

Tablet diameter 8 mm. Tablet shape: round, compound cup.

Example 14 Controlled Release Tablet Formulation Based on Eroding HPMCMatrix

HPMC added as part of the intragranular phase. Melt granulation

Tablet Composition:

Ingredient mg Tacrolimus 2 Lactose 80 PEG 6000 15 Poloxamer 188 6Metolose SH 90 80 Avicel PH200 60 Magnesium stearate 2 Total 245

The tablet formulation was based on melt granulation in a high shearmixer Pellmix ⅛. 16 g Micronized tacrolimus was mixed with 640 g lactose125 mesh and 120 g Polyethylene glycol 6000, 48 g Poloxamer 188 and 640g hydroxypropylmethylcellulose Metolose SH 90 15.000 cP in the highshear mixer. The jacket of the mixer bowl was heated to 80° C. and theblend was heated at an impeller rotation speed of 1000 rpm until meltingpoint of PEG and Poloxamer. After melting the kneading was continued for4 minutes at 800 rpm. The granulated was sieved through sieve size of0.7 mm and cooled on a tray. The granulate was mixed with 480 g AvicelPH200 for 3 minutes and for and after addition of 16 g magnesiumstearate for further 0.5 minute. The mixture was compressed into tabletson a single punch tabletting machine Diaf TM20. Tablet diameter 8 mm.Tablet shape: round, compound cup.

Example 15 Controlled Release Tablet Formulation Based on ErodingKollidon SR Matrix Added as Part of the Extragranular Phase

Tablet Composition:

Ingredient mg Tacrolimus 2 Lactose 80 Sodium lauryl sulfate 10 KollidonVA64 3 Lactose DC lac14 50 Kollidon SR 60 Magnesium stearate 1 Total 206

The tablet formulation was based on wet granulation in a high shearmixer Pellmix ⅛. 16 g Micronized tacrolimus was mixed with 640 g lactose125 mesh and 80 g natrium lauryl sulfate in the high shear mixer. A 15%aqueous solution of binder Kolllidon VA64 (Kollidon SR is a mixture ofpolyvinyl acetate and polyvinylpyrrolidon 80:20) was pumped to themixture at an impeller speed of 500 rpm at a feed rate of 20 g/min andsubsequently kneaded for 3 minutes. The granulate was dried in a traydryer and sieved through sieve size 0.7 mm.

The granulate was mixed with 400 g lactose DC Lac 14 and 480 g KollidonSR for 3 minutes and for and after addition of 8 g magnesium stearatefor further 0.5 minute. The mixture was compressed into tablets on asingle punch tabletting machine Diaf TM20. Tablet diameter: 8 mm. Tabletshape: round, compound cup.

Example 16 Enteric Coated Tablet Formulation Melt Granulation andEnteric-Coated Tablets

Tablet Composition:

Ingredient mg Tacrolimus 2 Lactose 80 PEG 6000 15 Poloxamer 188 6 AvicelPH200 60 Magnesium stearate 2 Total 165

The tablet formulation was based on melt granulation in a high shearmixer Pellmix ⅛. 16 g Micronized tacrolimus was mixed with 640 g lactose125 mesh and 120 g Polyethylene glycol 6000, 48 g Poloxamer 188 in thehigh shear mixer. The jacket of the mixer bowl was heated to 80° C. andthe blend was heated at a impeller rotation speed of 1000 rpm untilmelting point of PEG and Poloxamer. After melting the kneading wascontinued for 4 minutes at 800 rpm. The granulated was sieved throughsieve size of 0.7 mm and cooled on a tray. The granulate was mixed with480 g Avicel PH200 for 3 minutes and for and after addition of 16 gmagnesium stearate for further 0.5 minute. The mixture was compressedinto tablets on a single punch tabletting machine Diaf TM20. Tabletdiameter: 7 mm. Tablet shape: round, compound cup.

Enteric coating of the tablets is performed in accordance with theprocedure described in Example 11.

Example 17 In Vitro Dissolution Data

Compositions and dosage forms according to the previous examples weresubjected to in vitro dissolution tests using two different dissolutionmedia/tests.

-   -   A. Using the dissolution medium/test: 900 ml aqueous medium with        0.005% HPC (hydroxypropylcellulose) adjusted to pH=4.5 (USP        paddle method; rotation speed: 50 rpm), the following        dissolution profiles were found:

% Release Time Ex. 8 Ex. 9 - EC Ex. 10 (hours) Ex. 1 Ex. 3 Ex. 4 (Rsd %)(Rsd %) (Rsd %) 0 0 0 0 0 (0) 0 (0) 0 (0) 0.5 2 1 4 1.5 0 0 2 0 0 3 6 41 3 7.8 (11.1) 0.8 (32.3) 7.4 (9.8) 5 6 3 4 8 5 7 17 17.0 (8.3) 0.4(61.1) 13.3 (16.5) 10 20 14 15 40 32.2 (4.8) 11.0 (17.3) 36.0 (5.8) 1638 17 35.1 (9.6) 13.2 (12.1) 44.5 (5.4) 20 24 37

Dissolution profile for tablet cores of Example 9 in dissolution media:900 ml, aqueous media with 0.005% HPC (hydroxypropylcellulose) adjustedto pH=4.5. USP paddle method. Rotation speed: 50 rpm:

Time (minutes) % release Rsd % 0 0 0 5 27.2 15.1 10 49.1 10.9 20 80.78.0 35 98.9 5.4 42 102.7 3.6 52 104.9 2.0

Dissolution profile for enteric coated tablets example 9 in dissolutionmedium accord. to USP Method A, delayed release articles. USP Paddlemethod. Rotation speed: 50 rpm:

Time (minutes) % release Rsd % 0 0 NA 120 0 NA 155 84.8 12.8 165 102.9NA 175 101.0 3.5

Example 18 In Vivo Data Blood Plasma Concentration; Beagle Dogs

A. The following tacrolimus formulation was prepared:

Substance % mg Tacrolimus 0.89 1.00 HPMC Pharmacoat 606 20.37 22.81Lactose 200 mesh 20.37 22.81 Glyceryl monostearat Rylo MD50 58.38 65.38Total 100.00 112.00

Tacrolimus was dissolved in glyceryl monostearate at 80° C. The solutionwas sprayed onto a mixture of 100 g lactose and 100 ghydroxypropylmethylcellulose, Pharmacoat 606, in a fluid bed Strea-1 ata feed rate of 37 g/min. The resulting granular product was siftedthrough sieve nO. 0.7 mm and filled into hard gelatine capsules (112mg).

In vitro dissolution test (Dissolution media: 900 ml, aqueous mediaadjusted to pH4.5 with 0.005% HPC; USP paddle method; Rotation speed: 50rpm) of the formulation gave the following result:

Time (minutes) % release SD 0.5 83.6 21.6 1 93.6 7.14 2 97.1 8.98 4 97.47.77 8 98.8 7.74

An in vivo study of this formulation 0.5 mg in a Beagle dog, performedas described above under Methods, relative to Prograf®, 4×1 mg (Batchno.: 1C56050), gave the following results:

Blood concentrations (ng/mL) in dog no. F1183, after administration offormulation:

Formulation Time (hr) Prograf (4 mg) Invention dose adj. To 4 mg 0 0 00.5 0 0.7 1.0 5.8 4.2 1.5 16.6 10.15 2.0 13.7 14.0 3.0 5.1 10.85 4.0 3.39.1 6.0 2.4 5.6 8.0 2.3 4.2 12.0 2.3 3.15 24.0 1.2 2.1Relative bioavailability based on AUC (invention/Prograf): 151%.

B. The following tacrolimus formulation was prepared:

Substance % mg Tacrolimus 0.88 1.00 HPMC Pharmacoat 606 20.78 23.69Aeroperl 300 20.78 23.69 Glyceryl monostearate Rylo MD50 57.56 65.62Total 100.00 114.00

Tacrolimus was dissolved in glyceryl monostearate at 80° C. The solutionwas sprayed on a mixture of 100 g of Aeroperl® 300 (magnesium aluminiummetasilicate) and 100 g hydroxypropylmethylcellulose, Pharmacoat 606, ina fluid bed Strea-1 at a feed rate of 38 g/min. The granular product wassifted through sieve 0.7 mm and filled into hard gelatine capsules (114mg).

An in vivo study of this formulation 0.5 mg in a Beagle dog, performedas described above under Methods, relative to Prograf®, 4×1 mg (Batchno.: 1C56050), gave the following results:

Blood concentrations (ng/mL) in dog no. F1184, after administration offormulation:

Formulation Time (hr) Prograf (4 mg) Invention dose adj. to 4 mg 0 0 00.5 0.8 0.35 1.0 17.2 8.75 1.5 29.2 23.45 2.0 14.6 23.8 3.0 7.8 16.454.0 5.3 11.2 6.0 4.0 5.95 8.0 3.3 4.55 12.0 3.2 3.85 24.0 1.6 1.75Relative bioavailability based on AUC (invention/Prograf): 130%.

Example 19 In Vivo Data Blood Plasma Concentration; Göttingen Mini-Pigs

The following tacrolimus formulation was prepared:

Substance % mg Tacrolimus 0.52 1.00 HPMC Pharmacoat 606 12.02 22.83Lactose 200 mesh 12.02 22.83 Glyceryl monostearat Rylo MD50 34.44 65.44Magnesium stearate 1.00 1.90 Microcrystalline cellulose Avicel PH20040.00 76.00 Total 100.00 190.00

Tacrolimus was dissolved in glyceryl monostearate at 80° C. The solutionwas sprayed on a mixture of 100 g lactose and 100 ghydroxypropylmethylcellulose, Pharmacoat 606, in a fluid bed Strea-1 ata feed rate of 43 g/min. The granular product was sifted through sieve0.7 mm and mixed with 40% Avicel PH200 for 3 minutes in a Turbulablender and subsequently with 1% magnesium stearate for 0.5 minutes.Tablets of 190 mg were compressed on a single punch machine Diaf TM20.Tablet diameter 8 mm. Tablet shape: round, compound cup. Tablethardness: 42 N. Disintegration time: >55 min.

In vitro dissolution test (Dissolution media: 900 ml, aqueous mediaadjusted to pH4.5 with 0.005% HPC; USP paddle method; Rotation speed: 50rpm) of the formulation gave the following result:

Time (hours) Dissolution (%) SD 0.5 1.7 0.9 1 3.6 3.1 3 6.3 0.7 8 16.53.1 24 36.8 2.5

An in vivo study of this formulation 1 mg (assay 0.91 mg) in a femaleGöttingen mini-pig, performed as described above under Methods, relativeto Prograf® 1 mg (Batch no.: 1C5605D), gave the following results:

Blood concentrations (ng/mL) in pig no. 108003, after administration offormulation:

Formulation Time (hr) Prograf (1 mg) Invention (0.91 mg) 0 0.02 0 0.50.07 0.94 1.0 0.20 1.12 1.5 0.40 1.25 2.0 0.57 1.32 3.0 0.74 1.19 4.00.73 1.17 6.0 0.59 0.88 8.0 0.40 0.81 12.0 0.28 0.65 24.0 0.21 0.29 48.00.10 0.10Relative bioavailability based on AUC (invention/Prograf): 177%.

The invention claimed is:
 1. A method for suppressing a rejectionreaction by transplantation of an organ in a patient, the methodcomprising orally administering to the patient an extended releasepharmaceutical composition comprising a therapeutically effective amountof tacrolimus and one or more modifying release agents, wherein thecomposition releases at most 62% of the tacrolimus in the compositionwithin 15 hours when subjected to an in vitro dissolution test using USPPaddle method at a rotation speed of 50 rpm in a 900 mL aqueousdissolution medium with 0.005% hydroxypropylcellulose which has beenadjusted to pH 4.5.
 2. The method of claim 1, wherein the transplantedorgan is a kidney.
 3. The method of claim 1, wherein the transplantedorgan is a liver.
 4. The method of claim 1, wherein at most 60% w/w ofthe tacrolimus in the pharmaceutical composition is released within 15hours, when subjected to the in vitro dissolution test.
 5. The method ofclaim 1, wherein at most 50% w/w of the tacrolimus in the pharmaceuticalcomposition is released within 15 hours, when subjected to the in vitrodissolution test.
 6. The method of claim 1, wherein the pharmaceuticalcomposition provides an AUC_(fed)/AUC_(fasted) of at least 0.9.
 7. Themethod of claim 1, wherein the pharmaceutical composition provides pHindependent release.
 8. The method of claim 1, wherein the tacrolimus isdispersed or dissolved in a hydrophilic or water-miscible vehicle. 9.The method of claim 8, wherein the vehicle comprises poloxamer.
 10. Themethod of claim 8, wherein the vehicle comprises poloxamer andpolyethylene glycol.
 11. The method of claim 10, wherein thepolyethylene glycol and poloxamer are in a proportion by weight ofbetween about 1:3 and about 10:1.
 12. The method of claim 1, wherein thecomposition further comprises a solid carrier.
 13. The method of claim1, wherein the modifying release agent is a cellulose derivativeselected from hydroxypropyl methylcellulose (HPMC), hydroxypropylcellulose (HPC), methylcellulose, sodium carboxymethylcellulose,hydroxyethyl cellulose, poloxamers, polyoxyethylene stearates,poly-s-caprolactone, polyvinylpyrrolidone (PVP),polyvinylpyrrolidone-polyvinylacetate copolymer PVP-PVA, polymethacrylicpolymers, polyvinyl alcohol (PVA), poly(ethylene oxide) (PED), andmixtures thereof.
 14. The method of claim 1, wherein the compositioncomprises tacrolimus-containing particles.
 15. The method of claim 1,wherein the composition is in the form of a compressed tablet.
 16. Amethod for suppressing a rejection reaction by transplantation of anorgan in a patient, the method comprising orally administering to thepatient an extended release pharmaceutical composition comprising (a) atherapeutically effective amount of effective amount of tacrolimusdispersed or dissolved in a vehicle, and (b) one or more modifyingrelease agents, wherein (i) the vehicle comprises a poloxamer, and (ii)the composition releases at most 62% of the tacrolimus in thecomposition within 15 hours when subjected to an in vitro dissolutiontest using USP Paddle method at a rotation speed of 50 rpm in a 900 mLaqueous dissolution medium with 0.005% hydroxypropylcellulose which hasbeen adjusted to pH 4.5, which delays release of the tacrolimus to thedistal part of the duodenum thereby reducing tacrolimus inducedgastrointestinal side effects and metabolism of tacrolimus by CYP3A4.17. The method of claim 16, wherein the composition further comprises asolid carrier, and the vehicle further comprises polyethylene glycolhaving a molecular weight of from about 1,000 to about 35,000.